Russell RJ


Full name : Russell Robyn J

First name : Robyn J

Mail : Commonwealth Scientific and Industrial Research Organization (Australia) Ecosystems Science, Canberra, ACT 0200

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Country : Australia

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References (62)

Title : Isomer-specific comparisons of the hydrolysis of synthetic pyrethroids and their fluorogenic analogues by esterases from the cotton bollworm Helicoverpa armigera - Yuan_2015_Pestic.Biochem.Physiol_121_102
Author(s) : Yuan G , Li Y , Farnsworth CA , Coppin CW , Devonshire AL , Scott C , Russell RJ , Wu Y , Oakeshott JG
Ref : Pestic Biochem Physiol , 121 :102 , 2015
Abstract : The low aqueous solubility and chiral complexity of synthetic pyrethroids, together with large differences between isomers in their insecticidal potency, have hindered the development of meaningful assays of their metabolism and metabolic resistance to them. To overcome these problems, Shan and Hammock (2001) [7] therefore developed fluorogenic and more water-soluble analogues of all the individual isomers of the commonly used Type 2 pyrethroids, cypermethrin and fenvalerate. The analogues have now been used in several studies of esterase-based metabolism and metabolic resistance. Here we test the validity of these analogues by quantitatively comparing their hydrolysis by a battery of 22 heterologously expressed insect esterases with the hydrolysis of the corresponding pyrethroid isomers by these esterases in an HPLC assay recently developed by Teese et al. (2013) [14]. We find a strong, albeit not complete, correlation (r = 0.7) between rates for the two sets of substrates. The three most potent isomers tested were all relatively slowly degraded in both sets of data but three esterases previously associated with pyrethroid resistance in Helicoverpa armigera did not show higher activities for these isomers than did allelic enzymes derived from susceptible H. armigera. Given their amenability to continuous assays at low substrate concentrations in microplate format, and ready detection of product, we endorse the ongoing utility of the analogues in many metabolic studies of pyrethroids.
ESTHER : Yuan_2015_Pestic.Biochem.Physiol_121_102
PubMedSearch : Yuan_2015_Pestic.Biochem.Physiol_121_102
PubMedID: 26047117

Title : Functional screening of enzymes and bacteria for the dechlorination of hexachlorocyclohexane by a high-throughput colorimetric assay - Sharma_2014_Biodegradation_25_179
Author(s) : Sharma P , Jindal S , Bala K , Kumari K , Niharika N , Kaur J , Pandey G , Pandey R , Russell RJ , Oakeshott JG , Lal R
Ref : Biodegradation , 25 :179 , 2014
Abstract : Two distinct microbial dehalogenases are involved in the first steps of degradation of hexachlorocyclohexane (HCH) isomers. The enzymes, LinA and LinB, catalyze dehydrochlorination and dechlorination reactions of HCH respectively, each with distinct isomer specificities. The two enzymes hold great promise for use in the bioremediation of HCH residues in contaminated soils, although their kinetics and isomer specificities are currently limiting. Here we report the functional screening of a library of 700 LinA and LinB clones generated from soil DNA for improved dechlorination activity by means of a high throughput colorimetric assay. The assay relies upon visual colour change of phenol red in an aqueous medium, due to the pH drop associated with the dechlorination reactions. The assay is performed in a microplate format using intact cells, making it quick and simple to perform and it has high sensitivity, dynamic range and reproducibility. The method has been validated with quantitative gas chromatographic analysis of promising clones, revealing some novel variants of both enzymes with superior HCH degrading activities. Some sphingomonad isolates with potentially superior activities were also identified.
ESTHER : Sharma_2014_Biodegradation_25_179
PubMedSearch : Sharma_2014_Biodegradation_25_179
PubMedID: 23740574

Title : Identification of candidate odorant degrading gene\/enzyme systems in the antennal transcriptome of Drosophila melanogaster - Younus_2014_Insect.Biochem.Mol.Biol_53_30
Author(s) : Younus F , Chertemps T , Pearce SL , Pandey G , Bozzolan F , Coppin CW , Russell RJ , Maibeche-Coisne M , Oakeshott JG
Ref : Insect Biochemistry & Molecular Biology , 53 :30 , 2014
Abstract : The metabolism of volatile signal molecules by odorant degrading enzymes (ODEs) is crucial to the ongoing sensitivity and specificity of chemoreception in various insects, and a few specific esterases, cytochrome P450s, glutathione S-transferases (GSTs) and UDP-glycosyltransferases (UGTs) have previously been implicated in this process. Significant progress has been made in characterizing ODEs in Lepidoptera but very little is known about them in Diptera, including in Drosophila melanogaster, a major insect model. We have therefore carried out a transcriptomic analysis of the antennae of D. melanogaster in order to identify candidate ODEs. Virgin male and female and mated female antennal transcriptomes were determined by RNAseq. As with the Lepidoptera, we found that many esterases, cytochrome P450 enzymes, GSTs and UGTs are expressed in D. melanogaster antennae. As olfactory genes generally show selective expression in the antennae, a comparison to previously published transcriptomes for other tissues has been performed, showing preferential expression in the antennae for one esterase, JHEdup, one cytochrome P450, CYP308a1, and one GST, GSTE4. These largely uncharacterized enzymes are now prime candidates for ODE functions. JHEdup was expressed heterologously and found to have high catalytic activity against a chemically diverse group of known ester odorants for this species. This is a finding consistent with an ODE although it might suggest a general role in clearing several odorants rather than a specific role in clearing a particular odorant. Our findings do not preclude the possibility of odorant degrading functions for other antennally expressed esterases, P450s, GSTs and UGTs but, if so, they suggest that these enzymes also have additional functions in other tissues.
ESTHER : Younus_2014_Insect.Biochem.Mol.Biol_53_30
PubMedSearch : Younus_2014_Insect.Biochem.Mol.Biol_53_30
PubMedID: 25038463
Gene_locus related to this paper: drome-CG8424

Title : Draft Genome Sequence of Pandoraea sp. Strain SD6-2, Isolated from Lindane-Contaminated Australian Soil - Pushiri_2013_Genome.Announc_1_e00415
Author(s) : Pushiri H , Pearce SL , Oakeshott JG , Russell RJ , Pandey G
Ref : Genome Announc , 1 :e00415 , 2013
Abstract : Pandoraea sp. strain SD6-2 is a delta-hexachlorocyclohexane-degrading bacterial strain isolated from lindane-contaminated soil in Queensland, Australia. The genome of SD6-2 was sequenced to investigate its ability to degrade delta-hexachlorocyclohexane. Here we report the annotated genome sequence of this strain.
ESTHER : Pushiri_2013_Genome.Announc_1_e00415
PubMedSearch : Pushiri_2013_Genome.Announc_1_e00415
PubMedID: 23833132
Gene_locus related to this paper: 9burk-r7x7t2 , 9burk-r7x6f5 , 9burk-r7x6r1 , 9burk-r7wvd6 , 9burk-r7x694 , 9burk-r7x0f0 , 9burk-r7www6

Title : How many genetic options for evolving insecticide resistance in heliothine and spodopteran pests? - Oakeshott_2013_Pest.Manag.Sci_69_889
Author(s) : Oakeshott JG , Farnsworth CA , East PD , Scott C , Han Y , Wu Y , Russell RJ
Ref : Pest Manag Sci , 69 :889 , 2013
Abstract : The widely accepted paradigm for the development of insecticide resistance in field populations of insects is of selection for one or a very few genes of major effect. Limited genetic mapping data for organophosphate and pyrethroid resistance in heliothine and spodopteran pests generally agrees with this paradigm. However, other biochemical and transcriptomic data suggest a more complex set of changes in multiple P450 and esterase gene/enzyme systems in resistant strains of these species. We discuss possible explanations for this paradox, including the likely embedding of these genes in regulatory cascades and emerging evidence for their arrangement in large clusters of closely related genes. We conclude that there could indeed be an unusually large number of genetic options for evolving resistance in these species.
ESTHER : Oakeshott_2013_Pest.Manag.Sci_69_889
PubMedSearch : Oakeshott_2013_Pest.Manag.Sci_69_889
PubMedID: 23526801

Title : Heterologous expression and biochemical characterisation of fourteen esterases from Helicoverpa armigera - Teese_2013_PLoS.One_8_e65951
Author(s) : Teese MG , Farnsworth CA , Li Y , Coppin CW , Devonshire AL , Scott C , East P , Russell RJ , Oakeshott JG
Ref : PLoS ONE , 8 :e65951 , 2013
Abstract : Esterases have recurrently been implicated in insecticide resistance in Helicoverpa armigera but little is known about the underlying molecular mechanisms. We used a baculovirus system to express 14 of 30 full-length esterase genes so far identified from midgut cDNA libraries of this species. All 14 produced esterase isozymes after native PAGE and the isozymes for seven of them migrated to two regions of the gel previously associated with both organophosphate and pyrethroid resistance in various strains. Thirteen of the enzymes obtained in sufficient yield for further analysis all showed tight binding to organophosphates and low but measurable organophosphate hydrolase activity. However there was no clear difference in activity between the isozymes from regions associated with resistance and those from elsewhere in the zymogram, or between eight of the isozymes from a phylogenetic clade previously associated with resistance in proteomic and quantitative rtPCR experiments and five others not so associated. By contrast, the enzymes differed markedly in their activities against nine pyrethroid isomers and the enzymes with highest activity for the most insecticidal isomers were from regions of the gel and, in some cases, the phylogeny that had previously been associated with pyrethroid resistance. Phospholipase treatment confirmed predictions from sequence analysis that three of the isozymes were GPI anchored. This unusual feature among carboxylesterases has previously been suggested to underpin an association that some authors have noted between esterases and resistance to the Cry1Ac toxin from Bacillus thuringiensis. However these three isozymes did not migrate to the zymogram region previously associated with Cry1Ac resistance.
ESTHER : Teese_2013_PLoS.One_8_e65951
PubMedSearch : Teese_2013_PLoS.One_8_e65951
PubMedID: 23799064
Gene_locus related to this paper: helam-d5g3c9 , helam-s4wfz6

Title : Organophosphate and pyrethroid hydrolase activities of mutant Esterases from the cotton bollworm Helicoverpa armigera - Li_2013_PLoS.One_8_e77685
Author(s) : Li Y , Farnsworth CA , Coppin CW , Teese MG , Liu JW , Scott C , Zhang X , Russell RJ , Oakeshott JG
Ref : PLoS ONE , 8 :e77685 , 2013
Abstract : Two mutations have been found in five closely related insect esterases (from four higher Diptera and a hymenopteran) which each confer organophosphate (OP) hydrolase activity on the enzyme and OP resistance on the insect. One mutation converts a Glycine to an Aspartate, and the other converts a Tryptophan to a Leucine in the enzymes' active site. One of the dipteran enzymes with the Leucine mutation also shows enhanced activity against pyrethroids. Introduction of the two mutations in vitro into eight esterases from six other widely separated insect groups has also been reported to increase substantially the OP hydrolase activity of most of them. These data suggest that the two mutations could contribute to OP, and possibly pyrethroid, resistance in a variety of insects. We therefore introduced them in vitro into eight Helicoverpa armigera esterases from a clade that has already been implicated in OP and pyrethroid resistance. We found that they do not generally enhance either OP or pyrethroid hydrolysis in these esterases but the Aspartate mutation did increase OP hydrolysis in one enzyme by about 14 fold and the Leucine mutation caused a 4-6 fold increase in activity (more in one case) of another three against some of the most insecticidal isomers of fenvalerate and cypermethrin. The Aspartate enzyme and one of the Leucine enzymes occur in regions of the H. armigera esterase isozyme profile that have been previously implicated in OP and pyrethroid resistance, respectively.
ESTHER : Li_2013_PLoS.One_8_e77685
PubMedSearch : Li_2013_PLoS.One_8_e77685
PubMedID: 24204917
Gene_locus related to this paper: helam-d5g3c9 , helam-d5g3d2 , helam-d5g3d3 , helam-d5g3d4 , helam-d5g3d5 , helam-d5kx87 , helam-d5kx99 , helam-d5kxa9 , helam-d9iv61 , helam-d9iv62 , helam-h9zvh4 , helam-s4wfz6

Title : Structure and function of an insect alpha-carboxylesterase (alphaEsterase7) associated with insecticide resistance - Jackson_2013_Proc.Natl.Acad.Sci.U.S.A_110_10177
Author(s) : Jackson CJ , Liu JW , Carr PD , Younus F , Coppin C , Meirelles T , Lethier M , Pandey G , Ollis DL , Russell RJ , Weik M , Oakeshott JG
Ref : Proc Natl Acad Sci U S A , 110 :10177 , 2013
Abstract : Insect carboxylesterases from the alphaEsterase gene cluster, such as alphaE7 (also known as E3) from the Australian sheep blowfly Lucilia cuprina (LcalphaE7), play an important physiological role in lipid metabolism and are implicated in the detoxification of organophosphate (OP) insecticides. Despite the importance of OPs to agriculture and the spread of insect-borne diseases, the molecular basis for the ability of alpha-carboxylesterases to confer OP resistance to insects is poorly understood. In this work, we used laboratory evolution to increase the thermal stability of LcalphaE7, allowing its overexpression in Escherichia coli and structure determination. The crystal structure reveals a canonical alpha/beta-hydrolase fold that is very similar to the primary target of OPs (acetylcholinesterase) and a unique N-terminal alpha-helix that serves as a membrane anchor. Soaking of LcalphaE7 crystals in OPs led to the capture of a crystallographic snapshot of LcalphaE7 in its phosphorylated state, which allowed comparison with acetylcholinesterase and rationalization of its ability to protect insects against the effects of OPs. Finally, inspection of the active site of LcalphaE7 reveals an asymmetric and hydrophobic substrate binding cavity that is well-suited to fatty acid methyl esters, which are hydrolyzed by the enzyme with specificity constants ( approximately 10(6) M(-1) s(-1)) indicative of a natural substrate.
ESTHER : Jackson_2013_Proc.Natl.Acad.Sci.U.S.A_110_10177
PubMedSearch : Jackson_2013_Proc.Natl.Acad.Sci.U.S.A_110_10177
PubMedID: 23733941
Gene_locus related to this paper: luccu-E3aest7

Title : Draft Genome Sequence of Ralstonia sp. Strain GA3-3, Isolated from Australian Suburban Soil - Pearce_2013_Genome.Announc_1_e00414
Author(s) : Pearce SL , Pushiri H , Oakeshott JG , Russell RJ , Pandey G
Ref : Genome Announc , 1 : , 2013
Abstract : Ralstonia sp. strain GA3-3 is a hexachlorocyclohexane (HCH)-degrading bacterial strain isolated from suburban soil in Canberra, Australia. The genome of strain GA3-3 was sequenced to investigate its ability to degrade alpha-HCH. Here, we report the annotated genome sequence of this strain.
ESTHER : Pearce_2013_Genome.Announc_1_e00414
PubMedSearch : Pearce_2013_Genome.Announc_1_e00414
PubMedID: 23833131
Gene_locus related to this paper: alceu-catD1 , alceu-catD2 , cupnh-q0jyv4 , cupnh-q0k320 , cupnh-q0kd51 , ralpi-u3qr80 , cupnn-g0ewh7

Title : Testing the evolvability of an insect carboxylesterase for the detoxification of synthetic pyrethroid insecticides - Coppin_2012_Insect.Biochem.Mol.Biol_42_343
Author(s) : Coppin CW , Jackson CJ , Sutherland T , Hart PJ , Devonshire AL , Russell RJ , Oakeshott JG
Ref : Insect Biochemistry & Molecular Biology , 42 :343 , 2012
Abstract : Esterases have been implicated in metabolic resistance to synthetic pyrethroids in several insect species but little is yet known of the molecular basis for these effects. In this work modern directed evolution technology was used to test to what extent it is possible to genetically enhance the pyrethroid hydrolytic activity of the E3 carboxylesterase from the blowfly Lucilia cuprina. High throughput screening of a random mutant library with individual stereoisomers of fluorogenic analogues of two type II pyrethroids identified 17 promising variants that were then also tested with the commercial pyrethroid deltamethrin. Between them, these variants displayed significantly improved activities for all the substrates tested. Amino acid substitutions at ten different residues were clearly implicated in the improvements, although most only enhanced activity for a subset of the stereoisomers. Several new combinations of the most promising amino acid substitutions were then made, and negative epistatic effects were found in most of the combinations, but significant improvements were also found in a minority of them. The best mutant recovered contained three amino acid changes and hydrolysed deltamethrin at more than 100 times the rate of wild-type E3. Structural analysis shows that nine of the ten mutated residues improving pyrethroid or analogue activities cluster in putative substrate binding pockets in the active site, with the three mutations of largest effect all increasing the volume of the acyl pocket.
ESTHER : Coppin_2012_Insect.Biochem.Mol.Biol_42_343
PubMedSearch : Coppin_2012_Insect.Biochem.Mol.Biol_42_343
PubMedID: 22300675
Gene_locus related to this paper: luccu-E3aest7

Title : Genome sequence of the newly isolated chemolithoautotrophic Bradyrhizobiaceae strain SG-6C - Pearce_2011_J.Bacteriol_193_5057
Author(s) : Pearce SL , Pandey R , Dorrian SJ , Russell RJ , Oakeshott JG , Pandey G
Ref : Journal of Bacteriology , 193 :5057 , 2011
Abstract : Strain SG-6C (DSM 23264, CCM 7827) is a chemolithoautotrophic bacterium of the family Bradyrhizobiaceae. It can also grow heterotrophically under appropriate environmental conditions. Here we report the annotated genome sequence of this strain in a single 4.3-Mb circular scaffold.
ESTHER : Pearce_2011_J.Bacteriol_193_5057
PubMedSearch : Pearce_2011_J.Bacteriol_193_5057
PubMedID: 21742875

Title : Overexpressed esterases in a fenvalerate resistant strain of the cotton bollworm, Helicoverpa armigera - Wu_2011_Insect.Biochem.Mol.Biol_41_14
Author(s) : Wu S , Yang Y , Yuan G , Campbell PM , Teese MG , Russell RJ , Oakeshott JG , Wu Y
Ref : Insect Biochemistry & Molecular Biology , 41 :14 , 2011
Abstract : Enhanced detoxification is the major mechanism responsible for pyrethroid resistance in Chinese populations of Helicoverpa armigera. Previous work has shown that enhanced oxidation contributes to resistance in the fenvalerate-selected Chinese strain, YGF. The current study provides evidence that enhanced hydrolysis by esterase isozymes also contributes to the resistance in this strain. The average esterase activity of third instar YGF larvae was 1.9-fold compared with that of a susceptible SCD strain. Much of this difference was attributed to isozymes at two zones which hydrolysed the model carboxylester substrate 1-naphthyl acetate and also a 1-naphthyl analogue of fenvalerate. A preparation enriched for enzymes migrating to one of these zones from YGF was shown to hydrolyse fenvalerate with a specific activity of about 2.9 nmol/min/mg. This material was also matched by mass spectrometry with four putative carboxylesterase genes, all of which clustered within a phylogenetic clade of secreted midgut esterases. Quantitative PCR on these four genes showed several-fold greater expression in tissues of YGF compared to SCD but no differences was found in the number of copies of the genes between the strains.
ESTHER : Wu_2011_Insect.Biochem.Mol.Biol_41_14
PubMedSearch : Wu_2011_Insect.Biochem.Mol.Biol_41_14
PubMedID: 20875855
Gene_locus related to this paper: helam-h9zvh4

Title : Biochemical characterisation of MdCXE1, a carboxylesterase from apple that is expressed during fruit ripening - Souleyre_2011_Phytochemistry_72_564
Author(s) : Souleyre EJ , Marshall SD , Oakeshott JG , Russell RJ , Plummer KM , Newcomb RD
Ref : Phytochemistry , 72 :564 , 2011
Abstract : Esters are an important component of apple (Malusxdomestica) flavour. Their biosynthesis increases in response to the ripening hormone ethylene, but their metabolism by carboxylesterases (CXEs) is poorly understood. We have identified 16 members of the CXE multigene family from the commercial apple cultivar, 'Royal Gala', that contain all the conserved features associated with CXE members of the alpha/beta hydrolase fold superfamily. The expression of two genes, MdCXE1 and MdCXE16 was characterised in an apple fruit development series and in a transgenic line of 'Royal Gala' (AO3) that is unable to synthesise ethylene in fruit. In wild-type MdCXE1 is expressed at low levels during early stages of fruit development, rising to a peak of expression in apple fruit at harvest maturity. It is not significantly up-regulated by ethylene in the skin of AO3 fruit. MdCXE16 is expressed constitutively in wild-type throughout fruit development, and is up-regulated by ethylene in skin of AO3 fruit. Semi-purified recombinant MdCXE1 was able to hydrolyse a range of 4-methyl umbelliferyl ester substrates that included those containing acyl moieties that are found in esters produced by apple fruit. Kinetic characterisation of MdCXE1 revealed that the enzyme could be inhibited by organophosphates and that its ability to hydrolyse esters showed increasing affinity (K(m)) but decreasing turnover (k(cat)) as substrate acyl carbon length increases from C2 to C16. Our results suggest that MdCXE1 may have an impact on apple flavour through its ability to hydrolyse relevant flavour esters in ripe apple fruit.
ESTHER : Souleyre_2011_Phytochemistry_72_564
PubMedSearch : Souleyre_2011_Phytochemistry_72_564
PubMedID: 21315388
Gene_locus related to this paper: 9rosa-CXE1 , 9rosa-CXE2 , 9rosa-CXE3 , 9rosa-CXE4 , 9rosa-CXE5 , 9rosa-CXE6 , 9rosa-CXE7 , 9rosa-CXE8 , 9rosa-CXE9 , 9rosa-CXE10 , 9rosa-CXE11 , 9rosa-CXE12

Title : Cloning and biochemical characterization of a novel carbendazim (methyl-1H-benzimidazol-2-ylcarbamate)-hydrolyzing esterase from the newly isolated Nocardioides sp. strain SG-4G and its potential for use in enzymatic bioremediation - Pandey_2010_Appl.Environ.Microbiol_76_2940
Author(s) : Pandey G , Dorrian SJ , Russell RJ , Brearley C , Kotsonis S , Oakeshott JG
Ref : Applied Environmental Microbiology , 76 :2940 , 2010
Abstract : A highly efficient carbendazim (methyl-1H-benzimidazol-2-ylcarbamate, or MBC)-mineralizing bacterium was isolated from enrichment cultures originating from MBC-contaminated soil samples. This bacterium, Nocardioides sp. strain SG-4G, hydrolyzed MBC to 2-aminobenzimidazole, which in turn was converted to the previously unknown metabolite 2-hydroxybenzimidazole. The initial steps of this novel metabolic pathway were confirmed by growth and enzyme assays and liquid chromatography-mass spectrometry (LC-MS) studies. The enzyme responsible for carrying out the first step was purified and subjected to N-terminal and internal peptide sequencing. The cognate gene, named mheI (for MBC-hydrolyzing enzyme), was cloned using a reverse genetics approach. The MheI enzyme was found to be a serine hydrolase of 242 amino acid residues. Its nearest known relative is an uncharacterized hypothetical protein with only 40% amino acid identity to it. Codon optimized mheI was heterologously expressed in Escherichia coli, and the His-tagged enzyme was purified and biochemically characterized. The enzyme has a K(m) and k(cat) of 6.1 muM and 170 min(-1), respectively, for MBC. Radiation-killed, freeze-dried SG-4G cells showed strong and stable MBC detoxification activity suitable for use in enzymatic bioremediation applications.
ESTHER : Pandey_2010_Appl.Environ.Microbiol_76_2940
PubMedSearch : Pandey_2010_Appl.Environ.Microbiol_76_2940
PubMedID: 20228105
Gene_locus related to this paper: 9acto-c8cp46

Title : Gene identification and proteomic analysis of the esterases of the cotton bollworm, Helicoverpa armigera - Teese_2010_Insect.Biochem.Mol.Biol_40_1
Author(s) : Teese MG , Campbell PM , Scott C , Gordon KH , Southon A , Hovan D , Robin C , Russell RJ , Oakeshott JG
Ref : Insect Biochemistry & Molecular Biology , 40 :1 , 2010
Abstract : Some of the resistance of Helicoverpa armigera to conventional insecticides such as organophosphates and synthetic pyrethroids appears to be due to metabolic detoxification by carboxylesterases. To investigate the H. armigera carboxyl/cholinesterases, we created a data set of 39 putative paralogous H. armigera carboxyl/cholinesterase sequences from cDNA libraries and other sources. Phylogenetic analysis revealed a close relationship between these sequences and 70 carboxyl/cholinesterases from the recently sequenced genome of the silkworm, Bombyx mori, including several conserved clades of non-catalytic proteins. A juvenile hormone esterase candidate from H. armigera was identified, and B. mori orthologues were proposed for 31% of the sequences examined, however low similarity was found between lepidopteran sequences and esterases previously associated with insecticide resistance from other insect orders. A proteomic analysis of larval esterases then enabled us to match seven of the H. armigera carboxyl/cholinesterase sequences to specific esterase isozymes. All identified sequences were predicted to encode catalytically active carboxylesterases, including six proteins with N-terminal signal peptides and N-glycans, with two also containing C-terminal signals for glycosylphosphatidylinositol anchor attachment. Five of these sequences were matched to zones of activity on native PAGE at relative mobility values previously associated with insecticide resistance in this species.
ESTHER : Teese_2010_Insect.Biochem.Mol.Biol_40_1
PubMedSearch : Teese_2010_Insect.Biochem.Mol.Biol_40_1
PubMedID: 20005949
Gene_locus related to this paper: helam-d5g3c9 , helam-d5g3d2 , helam-d5g3d3 , helam-d5g3d4 , helam-d5g3d5 , helam-d5g3d9 , helam-d5g3e0 , helam-d5g3e1 , helam-d5g3e3 , helam-d5g3e4 , helam-d5g3e6 , helam-d5g3e7 , helam-d5g3e8 , helam-d5g3f1 , helam-d5g3f2 , helam-d5g3f3 , helam-d5g3f4 , helam-d5g3f9 , helam-d5g3g0 , helam-d5g3g1 , helam-d5g3g3 , helam-d5kx99 , helam-d5kxa9 , helam-d9iv61 , helam-f8rgs7 , helam-h9zvh4

Title : Esterase-based metabolic resistance to insecticides in heliothine and spodopteran pests - Farnsworth_2010_J.Pestic.Sci_35_275
Author(s) : Farnsworth CA , Teese MG , Yuan G , Li Y , Scott C , Zhang X , Wu Y , Russell RJ , Oakeshott JG
Ref : Journal of Pesticide Science , 35 :275 , 2010
Abstract : Elevated esterase activities and increased band intensities of multiple esterase isozymes after electrophoresis are commonly associated with resistance to organophosphate, pyrethroid and carbamate insecticides in various heliothine and spodopteran pests. One possible explanation for this involves a "master regulator" mutation in a more general chemical stress response. An association between elevated esterase activities and isozyme intensities has also been reported for resistance to the Cry1Ac toxin of Helicoverpa armigera. The basis for this is unclear albeit some involvement of esterases could be mediated by the toxin's affinity for N-acetyl galactosamine glycans on certain gut-expressed esterases in this species.
ESTHER : Farnsworth_2010_J.Pestic.Sci_35_275
PubMedSearch : Farnsworth_2010_J.Pestic.Sci_35_275

Title : Characterization of the phenylurea hydrolases A and B: founding members of a novel amidohydrolase subgroup - Khurana_2009_Biochem.J_418_431
Author(s) : Khurana JL , Jackson CJ , Scott C , Pandey G , Horne I , Russell RJ , Herlt A , Easton CJ , Oakeshott JG
Ref : Biochemical Journal , 418 :431 , 2009
Abstract : Mycobacterium brisbanense strain JK1, a bacterium capable of degrading the herbicide diuron, was isolated from herbicide-exposed soil. A gene/enzyme system with diuron hydrolase activity was isolated from this strain and named PUH (phenylurea hydrolase) B (puhB/PuhB) because of its close similarity to the previously characterized PUH A (puhA/PuhA). Both PUHs were heterologously expressed, purified and characterized. The PUHs were found to oligomerize as hexamers in solution, with each monomer containing a mononuclear Zn2+ active site. Sequence analysis showed that these enzymes belong to the metal-dependent amidohydrolase superfamily, although they contain a hitherto unreported Asn-X-His metal-binding motif and appear to form a novel sub-group within this superfamily. The effects of temperature and solvent on the enzymes were characterized. Determination of the kinetic parameters of the PUHs was used alongside Bronsted plots to develop a plausible catalytic mechanism, which is similar to that used by urease. In addition to the primary PUH activity, both enzymes are catalytically promiscuous, efficiently hydrolysing esters, carbamates and phosphotriesters. In fact, an analogue of diuron, in which the C-N bond was replaced by a C-O bond, was found to be turned over as efficiently as diuron, suggesting that the substrate specificity is predominantly determined by steric factors. The discovery of PuhA and PuhB on separate continents, and the absence of any other close homologues in the available sequence databases, poses a challenging question regarding the evolutionary origins of these enzymes.
ESTHER : Khurana_2009_Biochem.J_418_431
PubMedSearch : Khurana_2009_Biochem.J_418_431
PubMedID: 19000034
Gene_locus related to this paper: 9myco-b8r4k2

Title : Heterologous expression of the methyl carbamate-degrading hydrolase MCD - Naqvi_2009_J.Biotechnol_144_89
Author(s) : Naqvi T , Cheesman MJ , Williams MR , Campbell PM , Ahmed S , Russell RJ , Scott C , Oakeshott JG
Ref : J Biotechnol , 144 :89 , 2009
Abstract : The methyl carbamate-degrading hydrolase (MCD) of Achromobacter WM111 has considerable potential as a pesticide bioremediation agent. However this potential has been unrealisable until now because of an inability to express MCD in heterologous hosts such as Escherichia coli. Herein, we describe the first successful attempt to express appreciable quantities of MCD in active form in E. coli, and the subsequent characterisation of the heterologously expressed material. We find that the properties of this material closely match the previously reported properties of MCD produced from Achromobacter WM111. This includes the presence of two distinct forms of the enzyme that we show are most likely due to the presence of two functional translational start sites. The purified enzyme catalyses the hydrolysis of a carbamate (carbaryl), a carboxyl ester (alpha-naphthyl acetate) and a phophotriester (dimethyl umbelliferyl phosphate) and it is relatively resistant to thermal and solvent-mediated denaturation. The robust nature and catalytic promiscuity of MCD suggest that it could be exploited for various biotechnological applications.
ESTHER : Naqvi_2009_J.Biotechnol_144_89
PubMedSearch : Naqvi_2009_J.Biotechnol_144_89
PubMedID: 19770008

Title : Bridging the synaptic gap: neuroligins and neurexin I in Apis mellifera - Biswas_2008_PLoS.One_3_e3542
Author(s) : Biswas S , Russell RJ , Jackson CJ , Vidovic M , Ganeshina O , Oakeshott JG , Claudianos C
Ref : PLoS ONE , 3 :e3542 , 2008
Abstract : Vertebrate studies show neuroligins and neurexins are binding partners in a trans-synaptic cell adhesion complex, implicated in human autism and mental retardation disorders. Here we report a genetic analysis of homologous proteins in the honey bee. As in humans, the honeybee has five large (31-246 kb, up to 12 exons each) neuroligin genes, three of which are tightly clustered. RNA analysis of the neuroligin-3 gene reveals five alternatively spliced transcripts, generated through alternative use of exons encoding the cholinesterase-like domain. Whereas vertebrates have three neurexins the bee has just one gene named neurexin I (400 kb, 28 exons). However alternative isoforms of bee neurexin I are generated by differential use of 12 splice sites, mostly located in regions encoding LNS subdomains. Some of the splice variants of bee neurexin I resemble the vertebrate alpha- and beta-neurexins, albeit in vertebrates these forms are generated by alternative promoters. Novel splicing variations in the 3' region generate transcripts encoding alternative trans-membrane and PDZ domains. Another 3' splicing variation predicts soluble neurexin I isoforms. Neurexin I and neuroligin expression was found in brain tissue, with expression present throughout development, and in most cases significantly up-regulated in adults. Transcripts of neurexin I and one neuroligin tested were abundant in mushroom bodies, a higher order processing centre in the bee brain. We show neuroligins and neurexins comprise a highly conserved molecular system with likely similar functional roles in insects as vertebrates, and with scope in the honeybee to generate substantial functional diversity through alternative splicing. Our study provides important prerequisite data for using the bee as a model for vertebrate synaptic development.
ESTHER : Biswas_2008_PLoS.One_3_e3542
PubMedSearch : Biswas_2008_PLoS.One_3_e3542
PubMedID: 18974885
Gene_locus related to this paper: apime-b9vmq4 , apime-b9vmq5 , apime-b9vmq6 , apime-b9vmq7 , apime-a0a088atg1

Title : Jhe in Gryllus assimilis: cloning, sequence-activity associations and phylogeny - Crone_2007_Insect.Biochem.Mol.Biol_37_1359
Author(s) : Crone EJ , Zera AJ , Anand A , Oakeshott JG , Sutherland TD , Russell RJ , Harshman LG , Hoffmann FG , Claudianos C
Ref : Insect Biochemistry & Molecular Biology , 37 :1359 , 2007
Abstract : The 458 amino acid sequence of a mature JHE protein from the cricket Gryllus assimilis was identified after isolating the partial cDNA sequence encoding this protein from a fat body and midgut cDNA library. This hemimetabolan JHE sequence shows over 40% amino acid similarity to the known JHE sequences of several holometabolous insects. It also includes previously determined peptide sequences for G. assimilis JHE as well as two other motifs associated with JHE enzymes in holometabolous insects. The predicted molecular weight of the protein agrees with that of the JHE previously purified from G. assimilis. Partial genomic sequence encoding the Jhe contains two large (1330 and 2918bp) introns. No coding DNA sequence variation was observed over a 1293bp region between selected lines differing six to eight-fold in hemolymph JHE activity. However, a 19bp indel was found in one of the introns; the insertion was strongly associated with elevated hemolymph activity, both in the selected lines and in the F(2) progeny of crosses between them. Phylogenetic analyses localised the G. assimilis JHE to a clade containing dipteran and coleopteran JHEs, with lepidopteran JHEs occurring in a separate clade.
ESTHER : Crone_2007_Insect.Biochem.Mol.Biol_37_1359
PubMedSearch : Crone_2007_Insect.Biochem.Mol.Biol_37_1359
PubMedID: 17967354
Gene_locus related to this paper: 9orth-a5hsi6 , drome-CG8424

Title : High-resolution crystal structure of plant carboxylesterase AeCXE1, from Actinidia eriantha, and its complex with a high-affinity inhibitor paraoxon - Ileperuma_2007_Biochemistry_46_1851
Author(s) : Ileperuma NR , Marshall SD , Squire CJ , Baker HM , Oakeshott JG , Russell RJ , Plummer KM , Newcomb RD , Baker EN
Ref : Biochemistry , 46 :1851 , 2007
Abstract : Carboxylesterases (CXEs) are widely distributed in plants, where they have been implicated in roles that include plant defense, plant development, and secondary metabolism. We have cloned, overexpressed, purified, and crystallized a carboxylesterase from the kiwifruit species Actinidia eriantha (AeCXE1). The structure of AeCXE1 was determined by X-ray crystallography at 1.4 A resolution. The crystal structure revealed that AeCXE1 is a member of the alpha/beta-hydrolase fold superfamily, most closely related structurally to the hormone-sensitive lipase subgroup. The active site of the enzyme, located in an 11 A deep hydrophobic gorge, contains the conserved catalytic triad residues Ser169, Asp276, and His306. Kinetic analysis using artificial ester substrates showed that the enzyme can hydrolyze a range of carboxylester substrates with acyl groups ranging from C2 to C16, with a preference for butyryl moieties. This preference was supported by the discovery of a three-carbon acyl adduct bound to the active site Ser169 in the native structure. AeCXE1 was also found to be inhibited by organophosphates, with paraoxon (IC50 = 1.1 muM) a more potent inhibitor than dimethylchlorophosphate (DMCP; IC50 = 9.2 muM). The structure of AeCXE1 with paraoxon bound was determined at 2.3 A resolution and revealed that the inhibitor binds covalently to the catalytic serine residue, with virtually no change in the structure of the enzyme. The structural information for AeCXE1 provides a basis for addressing the wider functional roles of carboxylesterases in plants.
ESTHER : Ileperuma_2007_Biochemistry_46_1851
PubMedSearch : Ileperuma_2007_Biochemistry_46_1851
PubMedID: 17256879
Gene_locus related to this paper: actde-CXE1

Title : Hydrolysis of individual isomers of fluorogenic pyrethroid analogs by mutant carboxylesterases from Lucilia cuprina - Devonshire_2007_Insect.Biochem.Mol.Biol_37_891
Author(s) : Devonshire AL , Heidari R , Huang HZ , Hammock BD , Russell RJ , Oakeshott JG
Ref : Insect Biochemistry & Molecular Biology , 37 :891 , 2007
Abstract : We previously showed that wild-type E3 carboxylesterase of Lucilia cuprina has high activity against Type 1 pyrethroids but much less for the bulkier, alpha-cyano containing Type 2 pyrethroids. Both Types have at least two optical centres and, at least for the Type 1 compounds, we found that wild-type E3 strongly prefers the less insecticidal configurations of the acyl group. However, substitutions to smaller residues at two sites in the acyl pocket of the enzyme substantially increased overall activity, particularly for the more insecticidal isomers. Here we extend these analyses to Type 2 pyrethroids by using fluorogenic analogs of all the diastereomers of cypermethrin and fenvalerate. Wild-type E3 hydrolysed some of these appreciably, but, again, not those corresponding to the most insecticidal isomers. Mutations in the leaving group pocket or oxyanion hole were again generally neutral or deleterious. However, the two sets of mutants in the acyl pocket again improved activity for the more insecticidal acyl group arrangements as well as for the more insecticidal configuration of the cyano moiety on the leaving group. The activities of the best mutant enzyme against the analogs of the most insecticidal isomers of cypermethrin and fenvalerate were more than ten and a hundred fold higher, respectively, than those of wild-type. The implications for resistance development are discussed.
ESTHER : Devonshire_2007_Insect.Biochem.Mol.Biol_37_891
PubMedSearch : Devonshire_2007_Insect.Biochem.Mol.Biol_37_891
PubMedID: 17681228
Gene_locus related to this paper: luccu-E3aest7

Title : Only one esterase of Drosophila melanogaster is likely to degrade juvenile hormone in vivo - Crone_2007_Insect.Biochem.Mol.Biol_37_540
Author(s) : Crone EJ , Sutherland TD , Campbell PM , Coppin CW , Russell RJ , Oakeshott JG
Ref : Insect Biochemistry & Molecular Biology , 37 :540 , 2007
Abstract : Previously we identified juvenile hormone esterase (JHE) from Drosophila melanogaster by the criteria that it showed both appropriate developmental expression and kinetics for juvenile hormone (JH). We also noted three further esterases of D. melanogaster with some JHE-like characteristics, such as a GQSAG active site motif, a particular amphipathic helix, or close phylogenetic relationship with other JHEs. In this study, these JHE-like enzymes were expressed in vitro and their kinetic parameters compared with those of the previously identified JHE. Despite considerable phylogenetic distance between some of the esterases, they could all hydrolyse racemic JHIII. However, only the previously identified JHE had kinetic parameters (K(M) and k(cat)) towards various forms of JH (racemic or individual isomers of JHIII, JHII, JHI, and methyl farnesoate) consistent with a physiological role in JH regulation. Furthermore, only this JHE showed a preference for artificial substrates with acyl chain lengths similar to that of JH. This suggests that there is probably only one physiologically functional JHE in D. melanogaster but multiple esterases with JH esterase activity. Genomic comparisons of the selective JHE across 11 other Drosophila species showed a single orthologue in 10 of them but Drosophila willistoni has 16 full-length copies, five of them with the GQSAG motif and amphipathic helix.
ESTHER : Crone_2007_Insect.Biochem.Mol.Biol_37_540
PubMedSearch : Crone_2007_Insect.Biochem.Mol.Biol_37_540
PubMedID: 17517331
Gene_locus related to this paper: drome-CG8424

Title : Amplification of DNA from preserved specimens shows blowflies were preadapted for the rapid evolution of insecticide resistance - Hartley_2006_Proc.Natl.Acad.Sci.U.S.A_103_8757
Author(s) : Hartley CJ , Newcomb RD , Russell RJ , Yong CG , Stevens JR , Yeates DK , La Salle J , Oakeshott JG
Ref : Proc Natl Acad Sci U S A , 103 :8757 , 2006
Abstract : Mutations of esterase 3 confer two forms of organophosphate resistance on contemporary Australasian Lucilia cuprina. One form, called diazinon resistance, is slightly more effective against commonly used insecticides and is now more prevalent than the other form, called malathion resistance. We report here that the single amino acid replacement associated with diazinon resistance and two replacements associated with malathion resistance also occur in esterase 3 in the sibling species Lucilia sericata, suggesting convergent evolution around a finite set of resistance options. We also find parallels between the species in the geographic distributions of the polymorphisms: In both cases, the diazinon-resistance change is absent or rare outside Australasia where insecticide pressure is lower, whereas the changes associated with malathion resistance are widespread. Furthermore, PCR analysis of pinned specimens of Australasian L. cuprina collected before the release of organophosphate insecticides reveals no cases of the diazinon-resistance change but several cases of those associated with malathion resistance. Thus, the early outbreak of resistance in this species can be explained by the preexistence of mutant alleles encoding malathion resistance. The pinned specimen analysis also shows much higher genetic diversity at the locus before organophosphate use, suggesting that the subsequent sweep of diazinon resistance in Australasia has compromised the scope for the locus to respond further to the ongoing challenge of the insecticides.
ESTHER : Hartley_2006_Proc.Natl.Acad.Sci.U.S.A_103_8757
PubMedSearch : Hartley_2006_Proc.Natl.Acad.Sci.U.S.A_103_8757
PubMedID: 16723400

Title : Comparing the organophosphorus and carbamate insecticide resistance mutations in cholin- and carboxyl-esterases - Oakeshott_2005_Chem.Biol.Interact_157-158_269
Author(s) : Oakeshott JG , Devonshire AL , Claudianos C , Sutherland TD , Horne I , Campbell PM , Ollis DL , Russell RJ
Ref : Chemico-Biological Interactions , 157-158 :269 , 2005
Abstract : Mutant insect carboxyl/cholinesterases underlie over 60 cases of resistance to organophosphorus and/or carbamate insecticides. Biochemical and molecular data on about 20 of these show recurrent use of a very small number of mutational options to generate either target site or metabolic resistance. Moreover, the mutant enzymes are often kinetically inefficient and associated with significant fitness costs, due to impaired performance of the enzymes' original function. By contrast many bacterial enzymes are now known which can effectively detoxify these pesticides. It appears that the constraints of the genetic code and eukaryote genetic systems have severely limited the evolutionary response of insects to the widespread use of the insecticides over the last 60 years.
ESTHER : Oakeshott_2005_Chem.Biol.Interact_157-158_269
PubMedSearch : Oakeshott_2005_Chem.Biol.Interact_157-158_269
PubMedID: 16289012

Title : Hydrolysis of pyrethroids by carboxylesterases from Lucilia cuprina and Drosophila melanogaster with active sites modified by in vitro mutagenesis - Heidari_2005_Insect.Biochem.Mol.Biol_35_597
Author(s) : Heidari R , Devonshire AL , Campbell BE , Dorrian SJ , Oakeshott JG , Russell RJ
Ref : Insect Biochemistry & Molecular Biology , 35 :597 , 2005
Abstract : The cloned genes encoding carboxylesterase E3 in the blowfly Lucilia cuprina and its orthologue in Drosophila melanogaster were expressed in Sf9 cells transfected with recombinant baculovirus. Resistance of L. cuprina to organophosphorus insecticides is due to mutations in the E3 gene that enhance the enzyme's ability to hydrolyse insecticides. Previous in vitro mutagenesis and expression of these modifications (G137D, in the oxyanion hole and W251L, in the acyl pocket) have confirmed their functional significance. We have systematically substituted these and nearby amino acids by others expected to affect the hydrolysis of pyrethroid insecticides. Most mutations of G137 markedly decreased pyrethroid hydrolysis. W251L was the most effective of five substitutions at this position. It increased activity with trans permethrin 10-fold, and the more insecticidal cis permethrin >130-fold, thereby decreasing the trans:cis hydrolysis ratio to only 2, compared with >25 in the wild-type enzyme. Other mutations near the bottom of the catalytic cleft generally enhanced pyrethroid hydrolysis, the most effective being F309L, also in the presumptive acyl binding pocket, which enhanced trans permethrin hydrolysis even more than W251L. In these assays with racemic 1RS cis and 1RS trans permethrin, two phases were apparent, one being much faster suggesting preferential hydrolysis of one enantiomer in each pair as found previously with other esterases. Complementary assays with individual enantiomers of deltamethrin and the dibromo analogue of cis permethrin showed that the wild type and most mutants showed a marked preference for the least insecticidal 1S configuration, but this was reversed by the F309L substitution. The W251L/F309L double mutant was best overall in hydrolysing the most insecticidal 1R cis isomers. The results are discussed in relation to likely steric effects on enzyme-substrate interactions, cross-resistance between pyrethroids and malathion, and the potential for bioremediation of pyrethroid residues.
ESTHER : Heidari_2005_Insect.Biochem.Mol.Biol_35_597
PubMedSearch : Heidari_2005_Insect.Biochem.Mol.Biol_35_597
PubMedID: 15857765
Gene_locus related to this paper: drome-EST23aes07 , luccu-E3aest7

Title : Multiple mutations and gene duplications conferring organophosphorus insecticide resistance have been selected at the Rop-1 locus of the sheep blowfly, Lucilia cuprina - Newcomb_2005_J.Mol.Evol_60_207
Author(s) : Newcomb RD , Gleeson DM , Yong CG , Russell RJ , Oakeshott JG
Ref : Journal of Molecular Evolution , 60 :207 , 2005
Abstract : Sequences of the esterase gene alpha E7 were compared across 41 isogenic (IV) strains of the sheep blowfly, Lucilia cuprina, and one strain of the sibling species, L. sericata. The 1.2-kb region sequenced includes sites of two insecticide resistance mutations. Gly137Asp confers resistance to organophosphorus insecticides (OPs), particularly preferring diethyl OPs such as diazinon, while Trp251Leu prefers dimethyl OPs, and particularly malathion, with the additional presence of carboxylester moieties. We found that there are just eight haplotypes among the 41 chromosomes studied: two Gly137Asp containing haplotypes, two Trp251Leu containing haplotypes, and four susceptible haplotypes, including the L. sericata sequence. While phylogenetic analysis of these haplotypes suggests that the Asp137 and Leu251 mutations each arose at least twice, evidence for recombination was detected across the region, therefore single origins for these resistance mutations cannot be ruled out. Levels of linkage disequilibrium in the data are high and significant hitchhiking is indicated by Fay and Wu' s H test but not the Tajima test. A test of haplotype diversity indicates a paucity of diversity compared with neutral expectations. Both these results are consistent with a very recent selective sweep at the Lc alphaE7 locus. Interestingly, gene duplications of three different combinations of OP resistant haplotypes were identified in seven of the isogenic (IV) strains. All three types of duplication involve an Asp137 and a Trp251 haplotype. To examine whether more haplotypes existed before the hypothesised selective sweep, fragments of alpha E7 surrounding the resistance mutations were amplified from pinned material dating back to before OPs were used. Four new sequence haplotypes, not sampled in the survey of extant haplotypes, were obtained that are all associated with susceptibility. This is suggestive of a higher historical level of susceptible allelic diversity at this locus.
ESTHER : Newcomb_2005_J.Mol.Evol_60_207
PubMedSearch : Newcomb_2005_J.Mol.Evol_60_207
PubMedID: 15785849
Gene_locus related to this paper: luccu-E3aest7

Title : Two major classes of target site insensitivity mutations confer resistance to organophosphate and carbamate insecticides - Russell_2004_Pestic.Biochem.Physiol_79_84
Author(s) : Russell RJ , Claudianos C , Campbell PM , Horne I , Sutherland TD , Oakeshott JG
Ref : Pesticide Biochemistry and Physiology , 79 :84 , 2004
Abstract : Interspecific comparisons of bioassay and biochemical data suggest two major patterns of target site resistance to carbamates and organophosphates. Pattern I resistance, which is generally more effective for carbamates, has been shown in two sub-species of mosquitoes to be due to a particular Gly-Ser mutation in the oxyanion hole within the active site of one of their two acetylcholinesterase enzymes. Intriguingly, different substitutions at the equivalent site confer organophosphate hydrolytic ability on other esterases responsible for metabolic resistance in some other species. In the case of the aphid, Myzus persicae, Pattern I resistance is due to a Ser-Phe mutation in the vicinity of the acyl pocket of acetylcholinesterase. Pattern II resistance is at least as effective for organophosphates as it is for carbamates and may even be specific to organophosphates in some cases. Molecular studies on this pattern of resistance in three higher Diptera show that it is due to changes that constrict the acetylcholinesterase active site gorge and limit binding of the insecticide to the catalytic residues at the base of the gorge. One case of Pattern II resistance in the mosquito, Culex tritaeniorhynchus, involves the same site near the acyl pocket of acetylcholinesterase, albeit a different substitution, as that involved in Pattern I resistance in M. persicae.
ESTHER : Russell_2004_Pestic.Biochem.Physiol_79_84
PubMedSearch : Russell_2004_Pestic.Biochem.Physiol_79_84

Title : Hydrolysis of organophosphorus insecticides by in vitro modified carboxylesterase E3 from Lucilia cuprina - Heidari_2004_Insect.Biochem.Mol.Biol_34_353
Author(s) : Heidari R , Devonshire AL , Campbell BE , Bell KL , Dorrian SJ , Oakeshott JG , Russell RJ
Ref : Insect Biochemistry & Molecular Biology , 34 :353 , 2004
Abstract : Resistance of the blowfly, Lucilia cuprina, to organophosphorus (OP) insecticides is due to mutations in LcalphaE7, the gene encoding carboxylesterase E3, that enhance the enzyme's ability to hydrolyse insecticides. Two mutations occur naturally, G137D in the oxyanion hole of the esterase, and W251L in the acyl binding pocket. Previous in vitro mutagenesis and expression of these modifications to the cloned gene have confirmed their functional significance. G137D enhances hydrolysis of diethyl and dimethyl phosphates by 55- and 33-fold, respectively. W251L increases dimethyl phosphate hydrolysis similarly, but only 10-fold for the diethyl homolog; unlike G137D however, it also retains ability to hydrolyse carboxylesters in the leaving group of malathion (malathion carboxylesterase, MCE), conferring strong resistance to this compound. In the present work, we substituted these and nearby amino acids by others expected to affect the efficiency of the enzyme. Changing G137 to glutamate or histidine was less effective than aspartate in improving OP hydrolase activity and like G137D, it diminished MCE activity, primarily through increases in Km. Various substitutions of W251 to other smaller residues had a broadly similar effect to W251L on OP hydrolase and MCE activities, but at least two were quantitatively better in kinetic parameters relating to malathion resistance. One, W251G, which occurs naturally in a malathion resistant hymenopterous parasitoid, improved MCE activity more than 20-fold. Mutations at other sites near the bottom of the catalytic cleft generally diminished OP hydrolase and MCE activities but one, F309L, also yielded some improvements in OP hydrolase activities. The results are discussed in relation to likely steric effects on enzyme-substrate interactions and future evolution of this gene.
ESTHER : Heidari_2004_Insect.Biochem.Mol.Biol_34_353
PubMedSearch : Heidari_2004_Insect.Biochem.Mol.Biol_34_353
PubMedID: 15041019
Gene_locus related to this paper: luccu-E3aest7

Title : Enzymatic bioremediation: from enzyme discovery to applications - Sutherland_2004_Clin.Exp.Pharmacol.Physiol_31_817
Author(s) : Sutherland TD , Horne I , Weir KM , Coppin CW , Williams MR , Selleck M , Russell RJ , Oakeshott JG
Ref : Clinical & Experimental Pharmacology & Physiology , 31 :817 , 2004
Abstract : 1. Enzymatic bioremediation is potentially a rapid method of removing environmental pesticide residues. Applications include the treatment of residues resulting from agricultural production and processing industries, such as the treatment of irrigation waters, surface-contaminated fruit and vegetables and spent dip liquors. 2. A specific application for some organophosphate-degrading enzymes involves detoxification of nerve agent stockpiles. Effective and affordable remediation requires highly specialized enzymes, so protein engineering techniques are being used to improve properties of various source enzymes to enhance catalytic rates, stability and substrate range. 3. Trials with an optimized organophosphate-degrading enzyme have shown the feasibility of such technology in various applications. 4. The enzymes developed for environmental remediation for specific pesticide classes also have applications as antidotes for high-dose pesticide poisonings and as prophylaxis for people at risk of high pesticide doses.
ESTHER : Sutherland_2004_Clin.Exp.Pharmacol.Physiol_31_817
PubMedSearch : Sutherland_2004_Clin.Exp.Pharmacol.Physiol_31_817
PubMedID: 15566400

Title : Developmental expression and gene\/enzyme identifications in the alpha esterase gene cluster of Drosophila melanogaster - Campbell_2003_Insect.Mol.Biol_12_459
Author(s) : Campbell PM , de QRGC , Court LN , Dorrian SJ , Russell RJ , Oakeshott JG
Ref : Insect Molecular Biology , 12 :459 , 2003
Abstract : Here we show how the 10 genes of the alpha esterase cluster of Drosophila melanogaster have diverged substantially in their expression profiles. Together with previously described sequence divergence this suggests substantial functional diversification. By peptide mass fingerprinting and in vitro gene expression we have also shown that two of the genes encode the isozymes EST9 (formerly ESTC) and EST23. EST9 is the major 'alpha staining' esterase in zymograms of gut tissues in feeding stages while orthologues of EST23 confer resistance to organophosphorus insecticides in other higher Diptera. The results for EST9 and EST23 concur with previous suggestions that the products of the alpha esterase cluster function in digestion and detoxification of xenobiotic esters. However, many of the other genes in the cluster show developmental or tissue-specific expression that seems inconsistent with such roles. Furthermore, there is generally poor correspondence between the mRNA expression patterns of the remaining eight genes and isozymes previously characterized by standard techniques of electrophoresis and staining, suggesting that the alpha cluster might only account for a small minority of the esterase isozyme profile.
ESTHER : Campbell_2003_Insect.Mol.Biol_12_459
PubMedSearch : Campbell_2003_Insect.Mol.Biol_12_459
PubMedID: 12974951

Title : Kinetic efficiency of mutant carboxylesterases implicated in organophosphate insecticide resistance - Devonshire_2003_Pestic.Biochem.Physiol_76_1
Author(s) : Devonshire AL , Heidari R , Bell KL , Campbell PM , Campbell BE , Odgers WA , Oakeshott JG , Russell RJ
Ref : Pesticide Biochemistry and Physiology , 76 :1 , 2003
Abstract : Resistance to organophosphorus (OP) insecticides in Lucilia cuprina arises from two mutations in carboxylesterase E3 that enable it to hydrolyse the phosphate ester of various organophosphates, plus the carboxlyester in the leaving group in the case of malathion. These mutations are not found naturally in the orthologous EST23 enzyme in Drosophila melanogaster. We have introduced the two mutations (G137D and W251L) into cloned genes encoding E3 and EST23 from susceptible L. cuprina and D. melanogaster and expressed them in vitro with the baculovirus system. The ability of the resultant enzymes to hydrolyse the phosphate ester of diethyl and dimethyl organophosphates was studied by a novel fluorometric assay, which also provided a sensitive titration technique for the molar amount of esterase regardless of its ability to hydrolyse the fluorogenic substrate used. Malathion carboxylesterase activity was also measured. The G137D mutation markedly enhanced (>30-fold) hydrolysis of both classes of phosphate ester by E3 but only had a similar effect on the hydrolysis of dimethyl organophosphate in EST23. Introduction of the W251L mutation into either gene enhanced dimethyl (23-30-fold) more than diethyl (6-10-fold) organophosphate hydrolysis and slightly improved (2-4-fold) malathion carboxylesterase activity, but only at high substrate concentration.
ESTHER : Devonshire_2003_Pestic.Biochem.Physiol_76_1
PubMedSearch : Devonshire_2003_Pestic.Biochem.Physiol_76_1
Gene_locus related to this paper: luccu-E3aest7 , musdo-EST23aes07

Title : Evolution of an organophosphate-degrading enzyme: a comparison of natural and directed evolution - Yang_2003_Protein.Eng_16_135
Author(s) : Yang H , Carr PD , McLoughlin SY , Liu JW , Horne I , Qiu X , Jeffries CM , Russell RJ , Oakeshott JG , Ollis DL
Ref : Protein Engineering , 16 :135 , 2003
Abstract : Organophosphate-degrading enzyme from Agrobacterium radiobacter P230 (OPDA) is a recently discovered enzyme that degrades a broad range of organophosphates. It is very similar to OPH first isolated from Pseudomonas diminuta MG. Despite a high level of sequence identity, OPH and OPDA exhibit different substrate specificities. We report here the structure of OPDA and identify regions of the protein that are likely to give it a preference for substrates that have shorter alkyl substituents. Directed evolution was used to evolve a series of OPH mutants that had activities similar to those of OPDA. Mutants were selected for on the basis of their ability to degrade a number of substrates. The mutations tended to cluster in particular regions of the protein and in most cases, these regions were where OPH and OPDA had significant differences in their sequences.
ESTHER : Yang_2003_Protein.Eng_16_135
PubMedSearch : Yang_2003_Protein.Eng_16_135
PubMedID: 12676982

Title : The genomics of insecticide resistance - Oakeshott_2003_Genome.Biol_4_202
Author(s) : Oakeshott JG , Home I , Sutherland TD , Russell RJ
Ref : Genome Biol , 4 :202 , 2003
Abstract : Genomic technologies are revealing several mechanisms of insecticide resistance involving enhanced detoxification or reduced target-site sensitivity that had previously defied molecular analyses. Genome projects are also revealing some potentially far-reaching consequences for pest-insect genomes of the rapid accumulation of multiple resistance mutations in very short periods of evolutionary time.
ESTHER : Oakeshott_2003_Genome.Biol_4_202
PubMedSearch : Oakeshott_2003_Genome.Biol_4_202
PubMedID: 12540295

Title : Isolation of a Pseudomonas monteilli strain with a novel phosphotriesterase - Horne_2002_FEMS.Microbiol.Lett_206_51
Author(s) : Horne I , Harcourt RL , Sutherland TD , Russell RJ , Oakeshott JG
Ref : FEMS Microbiology Letters , 206 :51 , 2002
Abstract : A Pseudomonas monteilli strain designated C11 that uses the phosphotriester coroxon as its sole phosphorus source has been isolated Native PAGE and activity staining identified a single isozyme with significant phosphotriesterase activity in the soluble fraction of the cell This phosphotriesterase could hydrolyse both coumaphos and coroxon The hydrolysis product of coroxon diethylphosphate and the thion analogue coumaphos could not serve as phosphorus sources when added to the growth medium The majority of the phosphotriesterase and phosphatase activity was contained in the soluble fraction of the cell Phosphatase activity was inhibited by vanadate as well as by dialysis against the metal chelator EDTA Phosphotriesterase activity was not affected by either vanadate or dialysis with EDTA or 1,10-phenanthroline Phosphotriesterase activity was regulated by the amounts of both phosphate and coroxon in the medium whereas total phosphatase activity was regulated by phosphate but not coroxon A lack of hybridisation using a probe against the opd organophosphate degradation gene encoding a phosphotriesterase from Flavobacterium sp ATCC27551 against bulk DNA from P monteilli C11 suggested that this strain does not contain opd The work presented here indicates the presence of a novel phosphotriesterase in P monteilli C11
ESTHER : Horne_2002_FEMS.Microbiol.Lett_206_51
PubMedSearch : Horne_2002_FEMS.Microbiol.Lett_206_51
PubMedID: 11786256

Title : Development of a simple and sensitive fluorimetric method for isolation of coumaphos-hydrolysing bacteria - Harcourt_2002_Lett.Appl.Microbiol_34_263
Author(s) : Harcourt RL , Horne I , Sutherland TD , Hammock BD , Russell RJ , Oakeshott JG
Ref : Lett Appl Microbiol , 34 :263 , 2002
Abstract : AIMS: To develop a simple, rapid and sensitive fluorimetric assay to detect, isolate and characterize a soil bacterium capable of degrading the organophosphorus pesticide, coumaphos. METHODS AND
RESULTS: A high throughput microtitre plate-based method was used to quantify coumaphos hydrolysis by the bacterium. The fluorescent hydrolysis product of coumaphos, chlorferon, was detected at levels as low as 10 nmol l(-1). Incorporation of coumaphos into agar plates allowed the rapid detection of coumaphos-hydrolysing bacteria when exposed to an excitation wavelength of approximately 340 nm. The coumaphos-hydrolysing enzyme could be visualized when bacterial cell extracts were separated on SDS-PAGE, incubated with coumaphos and exposed to an excitation source as above.
CONCLUSIONS: This method is 100-fold more sensitive than the currently used spectrophotometric method for coumaphos. SIGNIFICANCE AND IMPACT OF THE STUDY: This is a unique and versatile tool to screen for bacteria possessing phosphotriesterase activity.
ESTHER : Harcourt_2002_Lett.Appl.Microbiol_34_263
PubMedSearch : Harcourt_2002_Lett.Appl.Microbiol_34_263
PubMedID: 11940156

Title : Identification of an opd (organophosphate degradation) gene in an Agrobacterium isolate - Horne_2002_Appl.Environ.Microbiol_68_3371
Author(s) : Horne I , Sutherland TD , Harcourt RL , Russell RJ , Oakeshott JG
Ref : Applied Environmental Microbiology , 68 :3371 , 2002
Abstract : We isolated a bacterial strain, Agrobacterium radiobacter P230, which can hydrolyze a wide range of organophosphate (OP) insecticides. A gene encoding a protein involved in OP hydrolysis was cloned from A. radiobacter P230 and sequenced. This gene (called opdA) had sequence similarity to opd, a gene previously shown to encode an OP-hydrolyzing enzyme in Flavobacterium sp. strain ATCC 27551 and Brevundimonas diminuta MG. Insertional mutation of the opdA gene produced a strain lacking the ability to hydrolyze OPs, suggesting that this is the only gene encoding an OP-hydrolyzing enzyme in A. radiobacter P230. The OPH and OpdA proteins, encoded by opd and opdA, respectively, were overexpressed and purified as maltose-binding proteins, and the maltose-binding protein moiety was cleaved and removed. Neither protein was able to hydrolyze the aliphatic OP malathion. The kinetics of the two proteins for diethyl OPs were comparable. For dimethyl OPs, OpdA had a higher k(cat) than OPH. It was also capable of hydrolyzing the dimethyl OPs phosmet and fenthion, which were not hydrolyzed at detectable levels by OPH.
ESTHER : Horne_2002_Appl.Environ.Microbiol_68_3371
PubMedSearch : Horne_2002_Appl.Environ.Microbiol_68_3371
PubMedID: 12089017

Title : Identification of a juvenile hormone esterase gene by matching its peptide mass fingerprint with a sequence from the Drosophila genome project - Campbell_2001_Insect.Biochem.Mol.Biol_31_513
Author(s) : Campbell PM , Harcourt RL , Crone EJ , Claudianos C , Hammock BD , Russell RJ , Oakeshott JG
Ref : Insect Biochemistry & Molecular Biology , 31 :513 , 2001
Abstract : Juvenile hormone esterase (JHE, EC from whole Drosophila melanogaster prepupae has previously been purified by selective precipitations, isoelectric focussing and two column chromatography steps. JHE bands from dried silver-stained SDS-PAGE gels of that material were digested with trypsin. The masses of the tryptic digest peptides were determined by MALDI-TOF mass spectrometry. Only one predicted gene product (CG8425) from the D. melanogaster genome matches the JHE tryptic fingerprint with high confidence. This predicted JHE sequence includes features that are conserved among all active members of the serine carboxylesterase multigene family as well as features peculiar to JHEs from other species. Also we show that this JHE can be purified by an alternative method using anion exchange chromotography followed by trifluoromethylketone affinity chromatography. A cDNA encoding this JHE was isolated using 3' and 5' RACE. This sequence is in agreement with the Drosophila genome project's prediction except that the sixth predicted intron is not removed; instead there is a stop codon followed by a polyadenylation signal and a polyA tail.
ESTHER : Campbell_2001_Insect.Biochem.Mol.Biol_31_513
PubMedSearch : Campbell_2001_Insect.Biochem.Mol.Biol_31_513
PubMedID: 11267890
Gene_locus related to this paper: drome-CG8424 , drome-CG8425

Title : MCE activities and malathion resistances in field populations of the australian sheep blowfly (Lucilia cuprina) - Smyth_2000_Heredity.(Edinb)_84 ( Pt 1)_63
Author(s) : Smyth KA , Boyce TM , Russell RJ , Oakeshott JG
Ref : Heredity (Edinb) , 84 ( Pt 1) :63 , 2000
Abstract : Malathion resistance has been shown to be the result of a single point mutation in the LcalphaE7 gene in four independently isolated chromosomes of Lucilia cuprina. The resultant amino acid substitution specifies high malathion carboxylesterase (MCE) activity. We have assayed MCE activities and resistance to malathion in three sets of field-derived samples, two sets of isogenic lines and five mass populations, and show that resistance to malathion in these samples is associated with high MCE activity in both sets of isogenic lines and four of the five mass populations. Additional mechanisms contributing to MCE activity or malathion resistance may be present in one of the mass populations. A second point mutation in LcalphaE7 is responsible for conferring diazinon resistance by encoding an increased organophosphate (OP) hydrolase activity. We also assayed diazinon resistances from the same three samples and show that diazinon and malathion resistances were in complete disequilibrium, with two exceptions. One exception involves the mass population with additional resistance mechanism(s) and the other involves three isogenic lines that are resistant to both insecticides. The molecular data for these lines suggest that they carry a duplication of the LcalphaE7 gene.
ESTHER : Smyth_2000_Heredity.(Edinb)_84 ( Pt 1)_63
PubMedSearch : Smyth_2000_Heredity.(Edinb)_84 ( Pt 1)_63
PubMedID: 10692012

Title : Reconstructing the diversification of alpha-esterases: comparing the gene clusters of Drosophila buzzatii and D. melanogaster - Robin_2000_J.Mol.Evol_51_149
Author(s) : Robin GC de Q , Claudianos C , Russell RJ , Oakeshott JG
Ref : Journal of Molecular Evolution , 51 :149 , 2000
Abstract : A cluster composed of 10 active alpha-esterase genes and a pseudogene is distributed over 60 kb in the Drosophila melanogaster genome. This paper describes the corresponding cluster in Drosophila buzzatii, whose lineage diverged from that of D. melanogaster when the subgenera Drosophila and Sophophora diverged about 50 Mya. With three exceptions we find that the composition of the cluster is conserved in the two lineages. The location of alpha E1 in D. melanogaster differs from that of its nearest relative in D. buzzatii, and alpha E4 has duplicated independently in the two lineages. The nature of these differences indicates that a mechanism exists whereby copies of genes can be placed in opposite orientation and nonadjacent positions within a gene cluster, although this does not seem to be a feature of earlier events in the cluster's evolution. The rates of amino acid change are not significantly different between orthologs, but the rates differ sevenfold among paralogs, indicating that very different selective forces are acting on the genes of the cluster. Mapping of sequence differences onto a model of the tertiary structure of the enzymes indicates that motifs contributing to substrate binding and catalysis have changed radically in the alphaE4s and suggest that this subgroup of alpha-esterases may be evolving into a substantially different functional niche.
ESTHER : Robin_2000_J.Mol.Evol_51_149
PubMedSearch : Robin_2000_J.Mol.Evol_51_149
PubMedID: 10948271
Gene_locus related to this paper: drobu-aes1a , drobu-aes02 , drobu-aes03 , drobu-aes4b , drobu-aes05

Title : The Evolution of an alpha-Esterase Pseudogene Inactivated in the Drosophila melanogaster Lineage - Robin_2000_Mol.Biol.Evol_17_563
Author(s) : Robin GC , Russell RJ , Cutler DJ , Oakeshott JG
Ref : Molecular Biology Evolution , 17 :563 , 2000
Abstract : Previous analyses of the alpha-esterase cluster of Drosophila melanogaster revealed 10 active genes and the DmalphaE4a-Psi pseudogene. Here, we reconstruct the evolution of the pseudogene from the sequences of 12 alleles from widely scattered D. melanogaster populations and single alleles from Drosophila simulans and Drosophila yakuba. All of the DmalphaE4a-Psi alleles contain numerous inactivating mutations, suggesting that pseudogene alleles are fixed in natural populations. Several lines of evidence also suggest that DmalphaE4a is now evolving without selective constraint in the D. melanogaster lineage. There are three polymorphic indels which result in frameshifts; a key nucleotide of the intron splice acceptor is polymorphic; the neutral mutation parameter is the same for replacement and silent sites; one of the nonsilent polymorphisms results in a stop codon; only 1 of the 13 replacement polymorphisms is biochemically conservative; residues that are conserved among active esterases have different states in DmalphaE4a-Psi; and there are about half as many transitional polymorphisms as transversional ones. In contrast, the D. simulans and D. yakuba orthologs DsalphaE4a and DyalphaE4a do not have the inactivating mutations of DmalphaE4a-Psi and appear to be evolving under the purifying selection typical of protein- encoding genes. For instance, there have been more substitutions in the introns than in the exons, and more in silent sites than in replacement sites. Furthermore, most of the amino acid substitutions that have occurred between DyalphaE4a and DsalphaE4a are located in sites that typically vary among active alpha-esterases rather than those that are usually conserved. We argue that the original alphaE4a gene had a function which it has lost since the divergence of the D. melanogaster and D. simulans lineages.
ESTHER : Robin_2000_Mol.Biol.Evol_17_563
PubMedSearch : Robin_2000_Mol.Biol.Evol_17_563
PubMedID: 10742048
Gene_locus related to this paper: drosi-aes04a , droya-aes04

Title : Poly(ethylene glycol) hydrogel-encapsulated fluorophore-enzyme conjugates for direct detection of organophosphorus neurotoxins - Russell_1999_Anal.Chem_71_4909
Author(s) : Russell RJ , Pishko MV , Simonian AL , Wild JR
Ref : Analytical Chemistry , 71 :4909 , 1999
Abstract : A simple approach is described for preparing poly-(ethylene glycol) hydrogel materials with encapsulated seminapthofluorescein (SNAFL)-organophosphorus hydrolase enzyme conjugates. Direct determination of enzyme-catalyzed neurotoxin hydrolysis is provided by the self-referencing, pH-sensitive dye SNAFL-1, whose emission spectrum changes at lambda = 550 in response to pH. Using spectrofluorimetry and paraoxon as a model organophosphate, paraoxon concentrations as low as 8 x 10(-7) M could be readily detected. On the basis of the signal-to-noise ratio, a detection limit of 16 nM was determined. The materials demonstrated high stability against enzyme-denaturing, leaching, and photobleaching when stored under ambient conditions.
ESTHER : Russell_1999_Anal.Chem_71_4909
PubMedSearch : Russell_1999_Anal.Chem_71_4909
PubMedID: 10565282

Title : Carboxyl\/cholinesterases: a case study of the evolution of a successful multigene family - Oakeshott_1999_Bioessays_21_1031
Author(s) : Oakeshott JG , Claudianos C , Russell RJ , Robin GC
Ref : Bioessays , 21 :1031 , 1999
Abstract : The evolution of organismal diversity among the Metazoa is dependent on the proliferation of genes and diversification of functions in multigene families. Here we analyse these processes for one highly successful family, the carboxyl/cholinesterases. One key to the expansion of the functional niche of this group of enzymes is associated with versatile substrate binding and catalytic machinery. Qualitatively new functions can be obtained by substitution of one or a very few amino acids. This crudely adapted new functionality is then refined rapidly by a pulse of change elsewhere in the molecule; in one case about 13% amino acid divergence occurred in 5-10 million years. Furthermore, we postulate that the versatility of the substrate binding motifs underpins the recruitment of several family members to additional noncatalytic signal transduction functions.
ESTHER : Oakeshott_1999_Bioessays_21_1031
PubMedSearch : Oakeshott_1999_Bioessays_21_1031
PubMedID: 10580988

Title : The same amino acid substitution in orthologous esterases confers organophosphate resistance on the house fly and a blowfly - Claudianos_1999_Insect.Biochem.Mol.Biol_29_675
Author(s) : Claudianos C , Russell RJ , Oakeshott JG
Ref : Insect Biochemistry & Molecular Biology , 29 :675 , 1999
Abstract : Organophosphate (OP) insecticide resistance in certain strains of Musca domestica is associated with reduction in the carboxylesterase activity of a particular esterase isozyme. This has been attributed to a 'mutant ali-esterase hypothesis', which invokes a structural mutation to an ali-esterase resulting in the loss of its carboxylesterase activity but acquisition of OP hydrolase activity. It has been shown that the mutation in Lucilia cuprina is a Gly137-->Asp substitution in the active site of an esterase encoded by the Lc alpha E7 gene (Newcomb, R.D., Campbell, P.M., Ollis, D.L., Cheah, E., Russell, R.J., Oakeshott, J.G., 1997. A single amino acid substitution converts a carboxylesterase to an organophosphate hydrolase and confers insecticide resistance on a blowfly. Proc. Natl. Acad. Sci. USA 94, 7464-7468). We now report the cloning and characterisation of the orthologous M. domestica Md alpha E7 gene, including the sequencing of cDNAs from the OP resistant Rutgers and OP susceptible sbo and WHO strains. The Md alpha E7 gene has the same intron structure as Lc alpha E7 and encodes a protein with 76% amino acid identity to Lc alpha E7. Comparisons between susceptible and resistance alleles show resistance in M. domestica is associated with the same Gly137-->Asp mutation as in L. cuprina. Bacterial expression of the Rutgers allele shows its product has OP hydrolase activity. The data indicate identical catalytic mechanisms have evolved in orthologous Md alpha E7 and Lc alpha E7 molecules to endow diazinon-type resistance on the two species of higher Diptera.
ESTHER : Claudianos_1999_Insect.Biochem.Mol.Biol_29_675
PubMedSearch : Claudianos_1999_Insect.Biochem.Mol.Biol_29_675
PubMedID: 10451921
Gene_locus related to this paper: musdo-EST23aes07

Title : Two different amino acid substitutions in the ali-esterase, E3, confer alternative types of organophosphorus insecticide resistance in the sheep blowfly, Lucilia cuprina - Campbell_1998_Insect.Biochem.Mol.Biol_28_139
Author(s) : Campbell PM , Newcomb RD , Russell RJ , Oakeshott JG
Ref : Insect Biochemistry & Molecular Biology , 28 :139 , 1998
Abstract : Two types of organophosphorus (OP) insecticide resistance are associated with reduced 'ali-esterase' (E3 isozyme) activity in Lucilia cuprina. The 'diazinon' resistance type shows generally greater resistance for diethyl than dimethyl OPs but no resistance to malathion. The 'malathion' resistance type shows generally greater resistance for dimethyl than diethyl OPs, low level diazinon resistance, but exceptionally high malathion resistance (600 x susceptible), the last being attributed to hydrolysis of the carboxylester groups which are peculiar to malathion (malathion carboxylesterase, MCE). E3 variants from diazinon resistant strains have previously been shown to have a Gly(137) --> Asp substitution that structural modelling predicts is only about 4.6 Angstrom from the gamma oxygen of the catalytic serine residue. Here we show that E3 variants from malathion resistant strains have a Trp(251) --> Leu substitution predicted to be about 4.3 Angstrom from that serine. We have expressed alleles of the gene encoding both resistance variants of E3 and an OP susceptible variant in a baculovirus system and compared the kinetics of their products. We find that both resistance substitutions reduce ali-esterase activity and enhance OP hydrolase activity. Furthermore the Gly(137) --, Asp substitution enhances OP hydrolase activity for a diethyl OP substrate (chlorfenvinphos) more than does the Trp(251) --> Leu substitution, which is consistent with the OP cross-resistance patterns. Trp(251) --> Leu also reduces the K-m for carboxylester hydrolysis of malathion about 10-fold to 21 mu M, which is consistent with increased RICE activity in malathion resistant strains. We then present a model in which the malathion carboxylesterase activity of the E3-Leu(251) enzyme is enhanced in vivo by its OP hydrolase activity. The latter activity enables it to reactivate after phosphorylation by malaoxon, the activated form of malathion, accounting for the exceptionally high level of resistance to malathion. We conclude that the two types of resistance can be explained by kinetic changes caused by the two allelic substitutions in the E3 enzyme
ESTHER : Campbell_1998_Insect.Biochem.Mol.Biol_28_139
PubMedSearch : Campbell_1998_Insect.Biochem.Mol.Biol_28_139
Gene_locus related to this paper: luccu-E3aest7

Title : Molecular Basis of Esterase-Mediated OP Resistance in Two Higher Diptera -
Author(s) : Oakeshott JG , Claudianos C , Campbell PM , Robin GC , Newcomb RD , Odgers WA , Harcourt RL , Russell RJ
Ref : In: Structure and Function of Cholinesterases and Related Proteins - Proceedings of Sixth International Meeting on Cholinesterases , (Doctor, B.P., Taylor, P., Quinn, D.M., Rotundo, R.L., Gentry, M.K. Eds) Plenum Publishing Corp. :497 , 1998

Title : A single amino acid substitution converts a carboxylesterase to an organophosphorus hydrolase and confers insecticide resistance on a blowfly - Newcomb_1997_Proc.Natl.Acad.Sci.U.S.A_94_7464
Author(s) : Newcomb RD , Campbell PM , Ollis DL , Cheah E , Russell RJ , Oakeshott JG
Ref : Proceedings of the National Academy of Sciences of the United States of America , 94 :7464 , 1997
Abstract : Resistance to organophosphorus (OP) insecticides is associated with decreased carboxylesterase activity in several insect species. It has been proposed that the resistance may be the result of a mutation in a carboxylesterase that simultaneously reduces its carboxylesterase activity and confers an OP hydrolase activity (the "mutant ali-esterase hypothesis"). In the sheep blowfly, Lucilia cuprina, the association is due to a change in a specific esterase isozyme, E3, which, in resistant flies, has a null phenotype on gels stained using standard carboxylesterase substrates. Here we show that an OP-resistant allele of the gene that encodes E3 differs at five amino acid replacement sites from a previously described OP-susceptible allele. Knowledge of the structure of a related enzyme (acetylcholinesterase) suggests that one of these substitutions (Gly137 --> Asp) lies within the active site of the enzyme. The occurrence of this substitution is completely correlated with resistance across 15 isogenic strains. In vitro expression of two natural and two synthetic chimeric alleles shows that the Asp137 substitution alone is responsible for both the loss of E3's carboxylesterase activity and the acquisition of a novel OP hydrolase activity. Modeling of Asp137 in the homologous position in acetylcholinesterase suggests that Asp137 may act as a base to orientate a water molecule in the appropriate position for hydrolysis of the phosphorylated enzyme intermediate.
ESTHER : Newcomb_1997_Proc.Natl.Acad.Sci.U.S.A_94_7464
PubMedSearch : Newcomb_1997_Proc.Natl.Acad.Sci.U.S.A_94_7464
PubMedID: 9207114
Gene_locus related to this paper: luccu-ACHE , luccu-E3aest7

Title : Biochemistry of esterases associated with organophosphate resistance in Lucilia cuprina with comparisons to putative orthologues in other Diptera - Campbell_1997_Biochem.Genet_35_17
Author(s) : Campbell PM , Trott JF , Claudianos C , Smyth KA , Russell RJ , Oakeshott JG
Ref : Biochemical Genetics , 35 :17 , 1997
Abstract : Esterase activities associated with organophosphate insecticide resistance in the Australian sheep blowfly, Lucilia cuprina, are compared with similar activities in other Diptera. The enzymes making the major contribution to methyl butyrate hydrolysis ("ali-esterase") in L. cuprina, M. domestica, and D. melanogaster comigrate during electrophoresis. The enzymes in L. cuprina and D. melanogaster correspond to the naphthyl acetate hydrolyzing E3 and EST23 isozymes of those species. These and previously published data suggest that the ali-esterases of all three species are orthologous. Strains of L. cuprina fall into four groups on the basis of quantitative determinations of their ali-estesterase, OP hydrolase, and malathion carboxylesterase activities and these groups correspond to their status with respect to two types of OP resistance. Strains susceptible to OP's have high ali-esterase, low OP hydrolase, and intermediate MCE activities; those resistant to malathion but not diazinon have low ali-esterase, intermediate OP hydrolase, and high MCE activities; those resistant to diazinon but not malathion have low ali-esterase, high OP hydrolase, and low MCE activities; those resistant to both OPs have low ali-esterase, high OP hydrolase, and high MCE activities. The correlated changes among the three biochemical and two resistance phenotypes suggest that they are all properties of one gene/enzyme system; three major allelic variants of that system explain OP susceptibility and the two types of OP resistance. Models are proposed to explain the joint contribution of OP hydrolase and MCE activities to malathion resistance and the invariant association of low ali-esterase and elevated OP hydrolase activities in either type of resistance.
ESTHER : Campbell_1997_Biochem.Genet_35_17
PubMedSearch : Campbell_1997_Biochem.Genet_35_17
PubMedID: 9238516
Gene_locus related to this paper: luccu-E3aest7

Title : Characterization of the EstP protein in Drosophila melanogaster and its conservation in drosophilids - Dumancic_1997_Biochem.Genet_35_251
Author(s) : Dumancic MM , Oakeshott JG , Russell RJ , Healy MJ
Ref : Biochemical Genetics , 35 :251 , 1997
Abstract : The beta-esterase cluster of D. melanogaster comprises two tandemly duplicated genes. Est6 encodes the well-characterized 5' gene, but the product of the second gene, denoted EstP, had not previously been identified. Here we show that the EstP gene encodes the carboxylesterase EST7. Expression of EstP using the Baculovirus system led to production of a carboxylesterase biochemically indistinguishable from EST7. Furthermore, a naturally occurring EstP variant produces greatly reduced amounts of EstP mRNA and no detectable EST7 protein. Finally, introduction of a wild-type copy of EstP by germline transformation into the variant strain confers the wild-type EST7 phenotype. We show that EST7 differs from EST6 in its substrate and inhibitor specificities and tissue distribution. Germline transformation experiments show that EstP expression is controlled by sequences located between 192 bp 5' and 609 bp 3' of the EstP coding region. Data comparisons with other drosophilid esterases suggest that the site of expression and hence the function, of EST7 has been conserved across lineages in both the subgenera Drosophila and Sophophora.
ESTHER : Dumancic_1997_Biochem.Genet_35_251
PubMedSearch : Dumancic_1997_Biochem.Genet_35_251
PubMedID: 9435945

Title : cDNA cloning, baculovirus-expression and kinetic properties of the esterase, E3, involved in organophosphorus resistance in Lucilia cuprina - Newcomb_1997_Insect.Biochem.Mol.Biol_27_15
Author(s) : Newcomb RD , Campbell PM , Russell RJ , Oakeshott JG
Ref : Insect Biochemistry & Molecular Biology , 27 :15 , 1997
Abstract : Resistance to organophosphorus insecticides (OPs) in the sheep blowfly, Lucilia cuprina, is associated with a non-staining phenotype of the carboxylesterase isozyme, E3 (E.C. Here, we show that a member of alpha-esterase multigene family, Lc alpha E7, encodes E3. An Lc alpha E7 cDNA has been isolated from an OP-susceptible strain and expressed in a baculovirus. The expressed product is the same as E3 in its electrophoretic mobility and preference for alpha-over beta-naphthyl acetate as substrate. Its preference (kcat/K(m)) for a range of carboxylester substrates is alpha-naphthyl butyrate > alpha-naphthyl propionate > alpha-naphthyl acetate > methylthiobutyrate > p-nitrophenyl acetate. The enzyme is potently inhibited by OPs (ki [paraoxon] = 6.3 +/- 1.4 x 10(7)/M/min, ki [chlorfenvinphos] = 5.9 +/- 0.6 x 10(7)/M/min) and exhibits a high turnover of methylthiobutyrate (1009/s), consistent with its proposed homology to the ali-esterase that is thought to mutate to confer OP resistance in Musca domestica. E3 shares 64% amino acid identity with its Drosophila melanogaster homologue, Dm alpha E7, and is also closely related to other esterases involved in OP resistance such as the B1 esterase of Culex pipiens (38%) and E4 of Myzus persicae (30%).
ESTHER : Newcomb_1997_Insect.Biochem.Mol.Biol_27_15
PubMedSearch : Newcomb_1997_Insect.Biochem.Mol.Biol_27_15
PubMedID: 9061925

Title : Comparison of an Esterase Associated with Organophosphate Resistance inLucilia cuprina with an Orthologue Not Associated with Resistance in Drosophila melanogaster - Parker_1996_Pestic.Biochem.Physiol_55_85
Author(s) : Parker AG , Campbell PM , Spackman ME , Russell RJ , Oakeshott JG
Ref : Pesticide Biochemistry and Physiology , 55 :85 , 1996
Abstract : Orthologous E3 and EST23 carboxylesterases have been enriched over 200-fold from organophosphate (OP) susceptible strains of Lucilia cuprina and Drosophila melanogaster, respectively. Mutants of E3 are associated with OP resistance but no resistance mutations of EST23 are known. The behaviours of the two enzymes were very similar during purification which involved differential centrifugation followed by three or four ion exchange and gel filtration chromatographic steps. Nondenaturing polyacrylamide gel electrophoresis and histochemical staining for esterase activity revealed no other esterases in the enriched material. Two-dimensional polyacrylamide gel electrophoresis (native followed by denaturing) showed that a major 70-kDa component of each preparation comigrates with E3 and EST23 activities, respectively. Kinetic properties of the enzymes are also very similar. Estimates of Km, Kcat, and Kcat/Km for alpha-naphthyl acetate are 42 +/- 18 μM, 19 sec-1, and 4.6 x 10(5) M-1 sec-1, respectively, for E3, and 62 +/- 25 μM, 23 sec-1, and 3.7 x 10(5) M-1 sec-1, for EST23. Both enzymes are potently inhibited by dibrom and less potently by another OP, diisopropylflurophosphate. E3 is also potently inhibited by paraoxon, whereas EST23 is at least 8-fold less susceptible to inhibition by paraoxon. This supports previous analyses of crude homogenates which showed that E3 is more susceptible to inhibition by paraoxon and fenitrooxon than is EST23 or the target site for OP action, acetylcholinesterase. It is proposed that the unusual affinity of E3 for such OPs is a necessary precondition for mutations that enable it to confer OP resistance on L. cuprina.
ESTHER : Parker_1996_Pestic.Biochem.Physiol_55_85
PubMedSearch : Parker_1996_Pestic.Biochem.Physiol_55_85
PubMedID: 8980033

Title : Isolation of alpha cluster esterase genes associated with organophosphate resistance in Lucilia cuprina - Newcomb_1996_Insect.Mol.Biol_5_211
Author(s) : Newcomb RD , East PD , Russell RJ , Oakeshott JG
Ref : Insect Molecular Biology , 5 :211 , 1996
Abstract : PCR primers designed from the alpha-esterase gene cluster of Drosophila melanogaster have been used to isolate fragments from eight esterase genes in the Australian sheep blowfly, Lucilia cuprina. Phylogenetic analysis suggests that three are homologues of the alpha E7, alpha E8 and alpha E9 genes of the alpha-esterase cluster of D. melanogaster. A further three are also probably alpha-esterases, whereas the remaining two more closely resemble beta-esterases. Transcripts for five of the alpha-esterase genes were detected by PCR in adult midgut, consistent with a role for these enzymes in digestion and/or detoxification. Based on the tissue distribution of these transcripts, Lc alpha E7 may possibly encode the esterase, E3, which is involved in organophosphate resistance.
ESTHER : Newcomb_1996_Insect.Mol.Biol_5_211
PubMedSearch : Newcomb_1996_Insect.Mol.Biol_5_211
PubMedID: 8799740
Gene_locus related to this paper: luccu-E3aest7

Title : Molecular cloning of an alpha-esterase gene cluster on chromosome 3r of Drosophila melanogaster - Russell_1996_Insect.Biochem.Mol.Biol_26_235
Author(s) : Russell RJ , Robin GC , Kostakos P , Newcomb RD , Boyce TM , Medveczky KM , Oakeshott JG
Ref : Insect Biochemistry & Molecular Biology , 26 :235 , 1996
Abstract : All or part of the alpha-esterase gene cluster in Drosophila melanogaster has been isolated by screening a YAC clone that spans cytological region 84D3-10 with consensus carboxyl/cholinesterase oligonucleotides. The cluster encompasses 11 putative esterase genes within 65 kb of genomic DNA and is one of the largest clusters of related protein-coding genes yet reported in Drosophila. The cluster must include the gene encoding the major alpha-esterase isozyme, EST9, which has previously been mapped to 84D3-5. It probably also includes the genes encoding the EST23, MCE and ALI esterases that have previously been mapped to 84D3-E2. The latter three are homologs of genes involved in organophosphate insecticide resistance in the sheep blowfly, Lucilia cuprina and the housefly, Musca domestica. Sequencing of one of the putative esterase genes in the Drosophila cluster, alpha E1, shows that it would encode features characteristic of an active carboxyl/cholinesterase, including the so-called catalytic triad, the nucleophilic elbow and oxyanion hole. It also shows that the closest relative of alpha E1 amongst previously published esterase sequences is ESTB1, which confers organophosphate resistance in Culex mosquitoes. We argue that we have cloned the D. melanogaster version of a major cluster of esterase genes which have variously mutated to confer organophosphate resistance in diverse Diptera.
ESTHER : Russell_1996_Insect.Biochem.Mol.Biol_26_235
PubMedSearch : Russell_1996_Insect.Biochem.Mol.Biol_26_235
PubMedID: 8900595
Gene_locus related to this paper: drome-aes01

Title : Duplication and divergence of the genes of the alpha-esterase cluster of Drosophila melanogaster - Robin_1996_J.Mol.Evol_43_241
Author(s) : Robin C , Russell RJ , Medveczky KM , Oakeshott JG
Ref : Journal of Molecular Evolution , 43 :241 , 1996
Abstract : The alpha-esterase cluster of D. melanogaster contains 11 esterase genes dispersed over 60 kb. Embedded in the cluster are two unrelated open reading frames that have sequence similarity with genes encoding ubiquitin-conjugating enzyme and tropomyosin. The esterase amino acid sequences show 37-66% identity with one another and all but one have all the motifs characteristic of functional members of the carboxyl/cholinesterase multigene family. The exception has several frameshift mutations and appears to be a pseudogene. Patterns of amino acid differences among cluster members in relation to generic models of carboxyl/cholinesterase protein structure are broadly similar to those among other carboxyl/cholinesterases sequenced to date. However the alpha-esterases differ from most other members of the family in: their lack of a signal peptide; the lack of conservation in cysteines involved in disulfide bridges; and in four indels, two of which occur in or adjacent to regions that align with proposed substrate-binding sites of other carboxyl/cholinesterases. Phylogenetic analyses clearly identify three simple gene duplication events within the cluster. The most recent event involved the pseudogene which is located in an intron of another esterase gene. However, relative rate tests suggest that the pseudogene remained functional after the duplication event and has become inactive relatively recently. The distribution of indels also suggests a deeper node in the gene phylogeny that separates six genes at the two ends of the cluster from a block of five in the middle.
ESTHER : Robin_1996_J.Mol.Evol_43_241
PubMedSearch : Robin_1996_J.Mol.Evol_43_241
PubMedID: 8703090
Gene_locus related to this paper: drome-aes01 , drome-aes02 , drome-aes03 , drome-aes04 , drome-aes05 , drome-aes06 , drome-aes08 , drome-aes09 , drome-aes10 , drome-EST23aes07

Title : Biochemical and Physiological Differences in the Malathion Carboxylesterase Activities of Malathion-Susceptible and -Resistant Lines of the Sheep Blowfly,Lucilia cuprina - Smyth_1996_Pestic.Biochem.Physiol_54_48
Author(s) : Smyth K-A , Walker VK , Russell RJ , Oakeshott JG
Ref : Pesticide Biochemistry and Physiology , 54 :48 , 1996
Abstract : Three malathion carboxylesterase (MCE) phenotypes have been described previously inLucilia cuprinaadults: low, intermediate, and high. The MCE specific activities of adults with the intermediate and high phenotypes are 21- and 33-fold higher than those with the low phenotype, but the high phenotype is almost 1000-fold more resistant to malathion than either of the other phenotypes. Here we show that MCE activity also peaks in Day 4 larvae from three lines representative of these phenotypes. The MCE activity of the low-line larvae is only 2.7- and 4-fold lower than those of the intermediate and high-line larvae, respectively. The relatively high MCE activity of the low-line larvae is largely explained by their crop activity. Assays in the presence of esterase inhibitors reveal three distinct types of MCE activity across the three lines and two developmental stages. Activity in the low-line adults is not completely inhibited by paraoxon and is classified as a subclass II carboxylesterase. The MCE activities in the larvae of the low and intermediate lines and adults from the intermediate line are inhibited by low concentrations of paraoxon, classifying them as subclass I carboxylesterases. The MCE activities in the high-line larvae and adults are also classified as subclass I carboxylesterases, but they are more sensitive to inhibition by triphenyl phosphate than those in the other two lines. These data suggest that MCE in the malathion-resistant high line may be structurally different from the MCEs in the malathion-susceptible intermediate and low lines.
ESTHER : Smyth_1996_Pestic.Biochem.Physiol_54_48
PubMedSearch : Smyth_1996_Pestic.Biochem.Physiol_54_48

Title : A cluster of esterase genes on chromosome 3R of Drosophila melanogaster includes homologues of esterase genes conferring insecticide resistance in Lucilia cuprina - Spackman_1994_Biochem.Genet_32_39
Author(s) : Spackman ME , Oakeshott JG , Smyth KA , Medveczky KM , Russell RJ
Ref : Biochemical Genetics , 32 :39 , 1994
Abstract : We identify an esterase isozyme in Drosophila melanogaster, EST 23, which shares biochemical, physiological, and genetic properties with esterase E3, which is involved in resistance to organophosphate insecticides in Lucilia cuprina. Like E3, the D. melanogaster EST 23 is a membrane-bound alpha-esterase which migrates slowly toward the anode at pH 6.8. Both enzymes have similar preferences for substrates with shorter acid side chain lengths. Furthermore, on the basis of their high sensitivity to inhibition by paraoxon and their insensitivity to inhibition by eserine sulfate, both enzymes were classified as subclass I carboxylesterases. The activity of each enzyme peaks early in development and, again, in the adult stage. Both enzymes are found in the male reproductive system and larval and adult digestive tissues, the latter being consistent with a role for these enzymes in organophosphate resistance. Fine structure deficiency mapping localized Est 23 to cytological region 84D3 to E1-2 on the right arm of chromosome 3. Moreover, we show that the genes encoding three other esterase phenotypes also map to the same region; these phenotypes involve allozymic differences in EST 9 (formerly EST C), ali-esterase activity, defined by the hydrolysis of methyl butyrate, and malathion carboxylesterase activity, defined by hydrolysis of the organophosphate malathion. This cluster corresponds closely to that encompassing E3 and malathion carboxylesterase on chromosome 4 in L. cuprina, the homologue of chromosome 3R in D. melanogaster.
ESTHER : Spackman_1994_Biochem.Genet_32_39
PubMedSearch : Spackman_1994_Biochem.Genet_32_39
PubMedID: 8031294

Title : A cluster of at least three esterase genes in Lucilia cuprina includes malathion carboxylesterase and two other esterases implicated in resistance to organophosphates - Smyth_1994_Biochem.Genet_32_437
Author(s) : Smyth KA , Russell RJ , Oakeshott JG
Ref : Biochemical Genetics , 32 :437 , 1994
Abstract : Three distinct malathion carboxylesterase (MCE) phenotypes have been identified among strains of Lucilia cuprina. The high-activity phenotype shows 1.6- and 33-fold more MCE specific activity than the intermediate- and low-activity phenotypes, respectively. Flies with high MCE activity are 1000-fold more resistant to malathion than flies with either low or intermediate MCE phenotypes, which are equally susceptible. High and low MCE specific activity are allelic and encoded by the Rmal gene on chromosome 4. Rmal is clustered within one map unit of two other esterase genes, Rop1 and E9, which are implicated in resistance to other organophosphate insecticides. Intermediate MCE specific activity is also inherited within the cluster, although its allelism to Rmal, Rop1, or E9 is unclear. The cluster does not contain the gene for the hemolymph esterase E4, which maps 6.1 map units from Rop1, on the other side of the bubbled wing marker. The cluster appears to be homologous to part of a tandem array of 11 esterase genes on chromosome 3R of Drosophila melanogaster.
ESTHER : Smyth_1994_Biochem.Genet_32_437
PubMedSearch : Smyth_1994_Biochem.Genet_32_437
PubMedID: 7748160

Title : Insecticide resistance and malathion carboxylesterase in the sheep blowfly, Lucilia cuprina - Whyard_1994_Biochem.Genet_32_9
Author(s) : Whyard S , Russell RJ , Walker VK
Ref : Biochemical Genetics , 32 :9 , 1994
Abstract : Resistance to the organophosphorus insecticide malathion in genetically related strains of the Australian sheep blowfly Lucilia cuprina was examined. Separate lines of blowflies were established by homozygosis of the fourth chromosome of the parental RM strain. Both the RM and the derived resistant (der-R) strains are approximately 100 times more resistant to malathion than the related susceptible der-S strain, resistance being correlated with a 45- to 50-fold increase in a malathion carboxylesterase (MCE) activity. MCE has a pH optimum ranging between 6.6 and 8.0 and is strongly inhibited by the carboxylesterase inhibitors triphenyl phosphate, paraoxon, and diisopropylfluorophosphate. Subcellular fractionation revealed that MCE was localized predominantly to the cytosol and mitochondria in both resistant and susceptible blowflies. A single MCE was purified to homogeneity from RM blowflies. It has a pI of 5.5, is a monomer of 60.5 kDa, and hydrolyzes malathion with a Vmax of 755 nmol/min/mg protein and a Km of 11.0 microM. L. cuprina have thus evolved a remarkable MCE which is faster and more efficient at hydrolyzing a specific insecticide than any other insect esterase vet described.
ESTHER : Whyard_1994_Biochem.Genet_32_9
PubMedSearch : Whyard_1994_Biochem.Genet_32_9
PubMedID: 8031296

Title : Evolutionary genetics of Drosophila esterases - Oakeshott_1993_Genetica_90_239
Author(s) : Oakeshott JG , van Papenrecht EA , Boyce TM , Healy MJ , Russell RJ
Ref : Genetica , 90 :239 , 1993
Abstract : Over 30 carboxylester hydrolases have been identified in D. melanogaster. Most are classified as acetyl, carboxyl or cholinesterases. Sequence similarities among most of the carboxyl and all the cholinesterases so far characterised from D. melanogaster and other eukaryotes justify recognition of a carboxyl/cholinesterase multigene family. This family shows minimal sequence similarities with other esterases but crystallographic data for a few non-drosophilid enzymes show that the family shares a distinctive overall structure with some other carboxyl and aryl esterases, so they are all put in one superfamily of/beta hydrolases. Fifteen esterase genes have been mapped in D. melanogaster and twelve are clustered at two chromosomal sites. The constitution of each cluster varies across Drosophila species but two carboxyl esterases in one cluster are sufficiently conserved that their homologues can be identified among enzymes conferring insecticide resistance in other Diptera. Sequence differences between two other esterases, the EST6 carboxyl esterase and acetylcholinesterase, have been interpreted against the consensus super-secondary structure for the carboxyl/cholinesterase multigene family; their sequence differences are widely dispersed across the structure and include substantial divergence in substrate binding sites and the active site gorge. This also applies when EST6 is compared across species where differences in its expression indicate a difference in function. However, comparisons within and among species where EST6 expression is conserved show that many aspects of the predicted super-secondary structure are tightly conserved. Two notable exceptions are a pair of polymorphisms in the substrate binding site of the enzyme in D. melanogaster. These polymorphisms are associated with differences in substrate interactions in vitro and demographic data indicate that the alternative forms are not selectively equivalent in vivo.
ESTHER : Oakeshott_1993_Genetica_90_239
PubMedSearch : Oakeshott_1993_Genetica_90_239
PubMedID: 8119594

Title : Biochemistry and physiology of esterases in organophosphate-susceptible and -resistant strains of the Australian sheep blowfly, Lucilia cuprina - Parker_1991_Pestic.Biochem.Physiol_41_305
Author(s) : Parker AG , Russell RJ , Delves AC , Oakeshott JG
Ref : Pesticide Biochemistry and Physiology , 41 :305 , 1991
Abstract : Eight esterases identified using native polyacrylamide gel electrophoresis were characterized in a susceptible and an organophosphate-resistant strain of Lucilia cuprina. Developmental profiles and tissue distributions were determined for all eight esterases and substrate and inhibitor specificities were also determined for six of them. Esterases E1 and E2 were isozymes of acetylcholinesterase, with E1 present throughout development and E2 largely confined to adults. Esterases E8, which was mainly confined to the neuromuscular and digestive tissues of early larvae, and E7, which was only recovered from 2- to 3-day-old adults, were not abundant enough for biochemical analysis. E4 and E13, classified as carboxylesterases, were only moderately sensitive to inhibition by organophosphates and found mainly in hemolymph. E3 and E9 were also identified as carboxylesterases but were highly sensitive to inhibition by organophosphates. They were the only enzymes found to differ between the two strains, both being detected only in the organophosphate-susceptible strain. In this strain they were both abundant in pupae, and E3 was also abundant in larvae and sexually mature adults, localized mainly in digestive tissues and the adult reproductive tract. The inhibitor, developmental, and tissue specificities of E3 all support genetic data reported previously that E3 is encoded by the major organophosphate resistance locus, ROP-1. While genetic data are not available for E9, its concentration in pupae suggests that it is unlikely to confer major gene resistance in feeding stages of L. cuprina.
ESTHER : Parker_1991_Pestic.Biochem.Physiol_41_305
PubMedSearch : Parker_1991_Pestic.Biochem.Physiol_41_305

Title : Molecular analysis of duplicated esterase genes in Drosophila melanogaster - Collet_1990_Mol.Biol.Evol_7_9
Author(s) : Collet C , Nielsen KM , Russell RJ , Karl M , Oakeshott JG , Richmond RC
Ref : Molecular Biology Evolution , 7 :9 , 1990
Abstract : Genomic clones containing sequences homologous to an esterase 6 (Est-6) cDNA clone were isolated from a library of Drosophila melanogaster DNA. Comparison of the genomic and cDNA sequences revealed that the Est-6 gene comprises two exons, one of 1,387 bp and one of 248 bp, separated by a short intron of 51 bp. Further sequencing revealed the presence of a tandem duplication of the Est-6 gene (denoted Est-P) which also has an exon of 1,387 bp and an exon of 248 bp, separated by a short intron of 56 bp. The two genes show similarities of 64% and 60% at the DNA and protein levels, respectively. The coding regions of the genes are 197 bases apart, and presumptive 5' regulatory sequences of Est-P overlap at least the 3' noncoding region of Est-6. Transcripts homologous to Est-P were detected in late larvae and adults of each sex, whereas Est-6 transcripts are present in all life stages but are predominant in adult males. This suggests different physiological functions for the products of the two genes. Southern and Northern blot hybridization analyses of the 20-kb region surrounding the Est-6/Est-P duplication failed to detect any other duplicated esterase genes, although this region is actively transcribed.
ESTHER : Collet_1990_Mol.Biol.Evol_7_9
PubMedSearch : Collet_1990_Mol.Biol.Evol_7_9
PubMedID: 2105433
Gene_locus related to this paper: drome-est6p , drome-este6

Title : Molecular cloning and characterization of esterase-6, a serine hydrolase of Drosophila - Oakeshott_1987_Proc.Natl.Acad.Sci.U.S.A_84_3359
Author(s) : Oakeshott JG , Collet C , Phillis RW , Nielsen KM , Russell RJ , Chambers GK , Ross V , Richmond RC
Ref : Proc Natl Acad Sci U S A , 84 :3359 , 1987
Abstract : The Est-6 gene of Drosophila melanogaster was cloned by screening libraries with synthetic oligonucleotides corresponding to tryptic peptides from purified esterase-6 (Est-6) protein. cDNA clones were isolated that hybridized in situ to the site of Est-6 on chromosome 3 at 69A1. Inserts in putative Est-6 cDNA clones were 1.85 kilobases (kb) long, and blot hybridization analysis of electrophoretically fractionated RNA, using a cDNA clone as a probe, revealed two transcripts, of 1.68 and 1.83 kb. The two transcripts showed the same developmental profile as the Est-6 protein. Neither transcript was detected in an Est-6-null line. The cDNA fragment was homologous to a 2.3-kb EcoRI-BamHI fragment in genomic clones, and this region was interrupted by the 8-kb B104 transposable element in the Est-6-null line. Conceptual translation of the cDNA sequence revealed a protein of 548 residues with 19% sequence similarity to acetylcholinesterase from the Torpedo ray.
ESTHER : Oakeshott_1987_Proc.Natl.Acad.Sci.U.S.A_84_3359
PubMedSearch : Oakeshott_1987_Proc.Natl.Acad.Sci.U.S.A_84_3359
PubMedID: 3106966
Gene_locus related to this paper: drome-este6